Closed photobioreactor system for continued daily in situ production of ethanol from genetically enhanced photosynthetic organisms with means for separation and removal of ethanol

ABSTRACT

The invention provides a device for growing genetically enhanced aquatic photoautotrophic organisms in a stable culture, causing said organisms to produce ethanol, and then separating, collecting, and removing the ethanol in situ.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a continuation-in-part application of the priorapplication to Woods et al., titled “Closed Photobioreactor System forContinued Daily In Situ Production, Separation, Collection, and Removalof Ethanol from Genetically Enhanced Microorganisms,” application Ser.No. 11/929,503, filed Oct. 30, 2007, now U.S. Pat. No. 7,682,821, whichin turn claims the benefit of U.S. Provisional Application No.60/864,091, filed Nov. 2, 2006. The text of both applications isincorporated by reference herein to the extent that there is noconsistency with the present disclosure.

BACKGROUND

The invention relates to the fields of engineering, microbiology, marinebiology, physical chemistry, and fluid dynamics.

The invention provides for an outdoor large volume closedphotobioreactor for the continued daily production of ethanol, or otherbiofuels, from a culture media comprising genetically enhancedcyanobacteria or algae and in situ separation of the ethanol from theculture media through evaporation by sunlight and subsequentcondensation and ethanol collection in the photobioreactor. Thephotobioreactor apparatus is designed to allow for the maintenance of ahigh density, stable culture comprising genetically enhancedcyanobacteria or algae and separation and collection of the ethanolproduced in the same apparatus. An embodiment of the invention is theremoval of ethanol from the culture comprising genetically enhancedcyanobacteria or algae, wherein the ethanol is removed from the culturewithout additional external manmade energy.

Given the high and escalating cost of fossil fuel based transportationfuels, the enormous world-wide demand for such fuels and the negativeenvironmental impact of the wide-spread use of these fuels, there hasbeen a significant market driven shift to the use of alternative fuelsthat are cleaner and renewable, namely biofuels. Currently, theproduction of biofuels, particularly ethanol, is dominated by theconversion of high cost feed substrates such as sugar cane, corn,rapeseed, palm oil and other terrestrial crops predominantly used asfood for human/animal consumption. While the technology exists toconvert these feedstocks to ethanol and biodiesel for use astransportation fuels, there is not sufficient arable land or fresh waterresources to meet the enormous demand of the global transportation fuelsmarket. The United States alone uses over 140 billion gallons ofgasoline for transportation fuel per year. The current U.S. output ofethanol made from corn is over 5 billion gallons annually. The economicimpact of the diversion of significant amounts of corn from thehuman/animal food market to the transportation fuels market has caused agreater than 50% increase in the market price of corn on globalcommodity markets. Such impacts on food commodity markets are notsustainable in the long-term, and large amounts of effort are beingexpended to find renewable alternatives that are cheaper and have thepotential for larger scale production.

The most predominant alternate technology being developed is biomassconversion, namely the conversion of cellulose based waste products tobiofuels using an industrial process. There remain significant technicalchallenges to bring this technology to a commercial reality. Given thehigh cost of the transportation of the cellulose feedstock to theprocessing facility and high capital costs, this technology could belimited in scale to facilities that can produce 5-100 million gallons ofbiofuel annually. Therefore, there is and will remain a need for anindustrial biofuels production technology that does not use or displacea feedstock that is for human/animal consumption, does not use arableland, can be made in very large quantities at a low price, and does notuse precious fresh water resources. One such technology is the use ofgenetically enhanced photoautotrophic cyanobacteria, algae, and otherphotoautotrophic organisms to convert internal sugars directly toethanol, butanol, pentanol and other higher alcohols and other biofuels.

For example, genetically modified cyanobacteria having constructscomprising DNA fragments encoding pyruvate decarboxylase (pdc) andalcohol dehydrogenase (adh) enzymes are described in U.S. Pat. No.6,699,696 (Woods, et al. for “Genetically modified cyanobacteria for theproduction of ethanol, the constructs and method thereof”).Cyanobacteria are photosynthetic bacteria which require light, inorganicelements, water, and a carbon source, generally carbon dioxide (CO₂), tometabolize and grow. The process using technology described in Woods, etal. has enabled the development of the industrial production of ethanolon a massive scale using readily available, cheap feed substrates,namely water and CO₂. The production of ethanol using geneticallyengineered cyanobacteria has also been described in PCT Published PatentApplication WO 2007/084477 (Fu et al. for “Methods and Compositions forEthanol Producing Cyanobacteria”).

The production of biofuels using genetically enhanced photoautotrophiccyanobacteria, algae, and other photoautotrophic organisms opens a newrealm in the industrial production of biofuels. The primary benefit ofthis technology is the combining of the process of the conversion ofsolar energy into cellular biochemical energy (the production ofinternal cellular “sugars”) with the fermentation of these internal“sugars” directly into ethanol in one single cell. This“direct-to-ethanol” approach eliminates the need to separately grow andharvest the feed substrate then convert it to the biofuel. Otherbenefits of such a technology are the ability to use non-arable,non-productive marginal or desert land for production facilities, theability to use saltwater, brackish water, fresh water or polluted wateras a feed substrate, the ability to recycle enormous amounts of carbondioxide into a transportation fuel and the ability to build massivescale production facilities with millions to billions of gallons ofannual production capacity, all based on a genetically enhancedphotoautotrophic organism.

Photoautotrophic organisms are those that can survive, grow andreproduce with energy derived entirely from the sun through the processof photosynthesis. Photosynthesis is essentially a carbon recyclingprocess through which inorganic carbon dioxide (CO₂) is combined withsolar energy, other nutrients and cellular biochemical processes tosynthesize carbohydrates and other compounds critical to life.Photosynthesis absorbs light in a limited range of the total solarspectrum, only in the wavelength range of 400-700 nm. This range onlyrepresents about half of the total solar energy. While at this timelittle can be done to expand the wavelength absorption range ofphotosynthesis, efforts can be made to optimize what energy can beabsorbed.

In the open environment, the overall photosynthetic efficiency rarelyexceeds 6%. A combination of factors including respiration during darkperiods, the length of the photoperiod, the intensity and incidence ofthe light, the chlorophyll content, available nutrients and stress allfurther reduce the efficiency of open plants in the natural outdoorenvironment. In laboratory photobioreactors, it is possible to achieve aphotosynthetic efficiency of greater than 24%. The goal of allphotobioreactor production systems is to optimize the environmentalconditions and fine tune the overall production process to achieve highbiomass production and photosynthesis yields well beyond those capablein the natural environment and in open pond growing systems.

Previous efforts for larger scale production have focused on growingphotoautotrophic organisms in open ponds or raceways that providesimilar growing conditions found in nature. A major drawback of thisapproach is that growing conditions cannot be well controlled, resultingin uncertain production outputs, batch contaminations and uncertainmanufacturing costs. These open systems are also not suited forefficiently cultivating the genetically enhanced (GE) organismsavailable today.

The current bottleneck for industrial photoautotrophic organismproduction is a lack of cost effective large-scale cultivation systemsutilizing photobioreactors. Very high volumetric production is necessaryto reduce the overall size of the installed production system as well asreduce the production and downstream processing cost. Key factors ofsuch systems are a high biomass concentration per volume, highphotosynthetic efficiency and the ability to have such systems use verylittle manmade energy. Designing cost effective, ultra large (millionsto billions of gallons of annual production output) manufacturingsystems that are needed to produce very large quantities of biofuels hasbeen a major unsolved technical challenge to date.

Various studies have resulted in designs of closed photobioreactors forculturing photoautotrophic organisms utilizing various technologies. Inthese controlled environments, much higher biomass productivity wasachieved, but the biomass growth rates were not high enough to offsetthe capital costs of the expensive systems utilized for the productionof low cost biofuels. Research in this field has focused on developingphotobioreactor systems of multiple designs including plate reactors(also known as flat panels), vertical gas-sparged photobioreactors,bubble column reactors, airlift reactors, external loop airlift reactorsand tubular photobioreactors. Each of these systems allow for varyingdegrees of process control and optimization, resulting in improvedgrowing conditions to achieve a predictable volume and cost. All ofthese systems have demonstrated the ability of higher volumetric biomassproduction when compared to open pond systems; however, all of thesesystems require significant external energy to operate the bioreactorsystems. For biofuels production, it will be necessary to limit theamount of energy required to operate the system to ensure the greatestpositive energy balance for the biofuels produced.

Photobioreactors are generally cylindrical or tubular in shape (pipe)(Yogev et al. in U.S. Pat. No. 5,958,761), are usually orientedhorizontal, and require additional energy to provide mixing (e.g.,pumps), thus adding significant capital and operational expense, theyhave no purposed airhead except that created by trapped O₂. Oxygen,produced by photosynthesis can also become trapped in these types ofsystems and negatively inhibit growth and biofuel production.Photobioreactors, such as bubble columns or airlifts, may be orientedvertically and agitated pneumatically which can reduce the need forfluid pumping. These bioreactors are primarily or solely for biomassaccumulation. Some photobioreactor designs rely on artificial lighting,e.g. fluorescent lamps, (such as described by Kodo et al. in U.S. Pat.No. 6,083,740). However, photobioreactors that do not utilize solarenergy, but instead rely solely on artificial light sources, require somuch energy input as to not be practical or cost effective forindustrial scale production of biofuels. Fu and Dexter (WO 2007/084477)used GE 6×26 watt bulbs to provide light to the Bioflo R® 110 bioreactorsystem.

Several studies of algae cultured in photobioreactors have usednarrow-bore tubes arranged in parallel and horizontal to the ground andon racks. These typically contain feed and harvest points to produce thebiomass and require large surface areas. These systems rely on churningprovided by pumping the biomass/growth medium through the piping atvarious high velocities. The cost of pumping in these systems willpreclude them from being used in the production of biofuels on a largescale. Such systems are also not practical on a very large scale such ascovering hundreds, if not thousands of hectares due to the high cost ofthe piping systems.

Bubble columns are typically translucent, large diameter verticallyoriented containers filled with algae suspended in liquid medium, inwhich gases are bubbled in at the bottom of the container. Sinceprecisely defined flow lines are not reproducibly formed in very largesystems, it can be difficult to control the mixing properties of thesystem, which can lead to low mass transfer coefficients, poorphotomodulation and low productivity.

Airlift reactors typically consist of vertically oriented concentrictubular containers, in which the gases are bubbled in at the bottom ofthe inner tube. The pressure gradient created at the bottom of the minortube creates an annular liquid flow upward through the inner tube andthen downward between the tubes. The external tube is made out oftranslucent material, while the inner tube is usually opaque. Therefore,the algae are exposed to light while passing between the tubes and todarkness while in the inner tube. The light-dark cycle is determined bythe geometrical design of the reactor (height, tube diameters) and byoperational parameters (e.g., gas flow rate).

Airlift bioreactors can have higher mass transfer coefficients and algalproductivity when compared to conventional mechanically stirred systems.Analogous to mammalian cell production, large bubbles results in poormass transfer of critical gases. Bubbles that are too small result ingreater shear near the point of bubble creation and, therefore, moredamaged or killed cells. Both damaged and killed cells can releasecomponents into the growth medium, that if too high, can greatly impactthe health and thus the productivity of the system. However, controlover the flow patterns within a very large airlift bioreactor to achievea desired level of mixing and photomodulation is difficult orimpractical. The energy requirement for an airlift photobioreactor istypically much lower than that for a stirred system and may be suitablefor higher value products than commodity transportation fuel, but eventhe pumping costs required for an airlift photobioreactor are too greatfor low value commodity transportation fuels.

Moreover, because of geometric design constraints in most currentsystems, cylindrical-photobioreactors suffer from low productivity whenused for large-scale outdoor algae production, due to factors related tolight reflection and auto-shading effects (in which one column isshading the other). This technology is impractical for use in producinglow value commodity transportation fuels such as ethanol.

It is important for optimum facility design and engineering tounderstand that when growing photosynthetic organisms at high density,shading of cells by other cells will reduce overall solar absorption.

Mixing mechanisms present a challenge in a bulk bioreactor and can beproblematic once the cells pass from the mixing area of the bioreactorto the solar collection tubes where photosynthesis occurs. A majorchallenge to scale-up in a photobioreactor systems is increased shearstress from mixing or turbulence that results in cell damage (Gudin C.,Dhaumont D. 1991. “Cell fragility is a key problem of microalgae massproduction in closed photobioreactors,” Bioresource Technology38:145-151). Cells are often more resistant to static hydrodynamic shearand less resistant to shear created by a liquid/air surface. Cell damageand lysis can occur at several points, including bubble creation, bubblerising and, as for mammalian cells, bubbles bursting at the liquid/airinterface. Plant cell walls often contain cellulosic material that givethem high tensile strength, but may have extremely low shear resistance.The fixed blade impellers or excessive airflows in airlift bioreactorsproduce high shear rates that result in cell breakage. The optimum levelof turbulence for mixing, which creates shear stress for cells, is aresult of fluid flow and gas velocity. As the cell density increases,the viscosity of the fluid rises, which works against uniform mixing andsubsequent optimum mass transfer of nutrients. High airflow rates athigh cell densities in an airlift bioreactor can result in shearbecoming too great and cell breakage occurring. Algae, similar to otherspecies of plants in suspension culture, vary in the resistance toshear. This has been a major challenge in developing a standardphotobioreactor in which all cells can be grown.

Rapid alteration between high light intensities and darkness haveconsistently been shown to significantly enhance the efficiency ofphotosynthesis, with shorter cycles having greater effects (Matthijs,et. al. Application of light emitting diodes in bioreactors: flashinglight effects and energy economy in algal culture. Biotechnol. Bioeng.50:98-107). It has been speculated that the reduction of electronacceptors in photosystem II (PSII), with the corresponding oxidation ofthose acceptors in the dark, results in high solar energy capture duringthe light. A solar absorption tube system containing clear and darkareas with in-line mixers and an optimized residence time through fluidflow control should be capable of light to dark cycles from fractions ofa minute to several minutes as dictated by the species of algae. Abetter way to get light dark transitions is to mix algae so they arealternately shaded by other algae or exposed to light. The overallefficiency of the system depends on its area and its output. A dark areais not contributing to output. The algae may be more efficient, but thesystem as a whole is not. But if shading by algae is used, light isalways being absorbed by an active element. Janssen, M, et. al.(Scale-up aspects of photobioreactor s: effects of mixing inducedlight/dark cycles. J. Appl. Phycol. 12:225-237.) demonstrated that alight gradient, which occurs in natural sunlight over a typical day, hasa significant impact on biomass yield from a given energy of light. Itis expected that as one moves away from the equator, either north orsouth, that the light gradient will be more pronounced over the year,resulting in lower efficiencies when moving away from the equator. Thedesign of many air-lift photobioreactors results in lower than expectedbiomass, even when carbon dioxide and other nutrients are increased.This is due to the physical design of most systems that facilitatesmedium length light/dark cycles, which have been shown to reduce biomassyield. Churning is too rapid in fast-moving airlift systems and bubblecolumn systems.

Airlift systems can be designed to provide optimal mass transfer ofoxygen and carbon dioxide, although at dry weight densities over 70 g/Lmaintaining adequate dissolved carbon dioxide becomes difficult. At highcell densities and during high rates of photosynthesis, the productionof oxygen and the rapid utilization of carbon dioxide often requiredventing of the system. This becomes an issue in long sealed phototubesand is a primary reason that external loop phototubes have limitedlengths when not vented. But these pipe or small diameter tube systemsare predominantly for biomass production and ethanol as a directlyproduced product is not being made.

As early as 1959, Tulecke and Nickell, and in 1963 by Wang and Staba,produced 20 liter bioreactors for the culture of plant cells. In theearly 1970's, Kato and his colleagues at the Japan Tobacco and SaltPublic Corporation investigated the use of air in mixing up to 1500liters. Later, a 20,000 liter system was used by Noguchi at the samecorporation. In the mid-1980's, Wagner and Vogelmann demonstrated thatthe airlift system was superior to all others in providing goodproductivity and a well defined and controlled system of parameters,resulting in reproducible flow characteristics. They further suggestedthat fluid movement can be better controlled through the use of aninternal draft tube through which the air mixture is bubbled. Althoughsuch systems can be scaled up significantly, their operation requiresmuch external energy and they are therefore not cost effective for useas production systems for biofuels. In addition, mutual shading invertical airlift systems has an impact on the total installed systemcapacity in a given footprint.

Javanmardian and Palsson (1991, High-density photoautotrophic algalcultures: design, construction, and operation of a novel photobioreactorsystem. Biotechnology & Bioengineering: 38, p 1182-1189,) developed anequation for the depth of light penetration. Applying this equation to ahigh density pond system suggests that at cell densities of 50 g/L,light penetration would be less than 2 mm. This demonstrates that lightpenetration clearly limits algal biomass production in typical open pondsituations. Ogbanna and Tanaka (1997, Industrial-size photobioreactor s.Chemtech: 27(7), p 43-49.) demonstrated that photosynthesis ismaintained with a light intensity of 7.3 μmol/m2/s. In addition, Lee andPalsson (1994, High-density algal photobioreactors using light-emittingDiodes. Biotechnology and Bioengineering: 44, p 1161-1167,) found thatboth light path and light intensity increased algal biomass productionwith light path possibly having a greater impact.

People have worked on ways to supplement natural light with artificiallighting in order to increase the efficiency of photosynthesis. Lee andPalsson (High density algal photobioreactors using light emittingdiodes. Biotech. BioEng. Vol 44, 1161-1167:1994) used highly efficientlight-emitting diodes (LED comprising gallium aluminum arsenide chips)to demonstrate that artificial light at specific wavelengths (680 nmmonochromatic red light) could significantly increase the density of thecell culture. They found that supplementation with light at a wavelengthof 680 nm would produce a cell concentration of more than 2×109 cells/ml(or more than 6.6% v/v) and an oxygen production rate as high as 10 mmoloxygen/L culture/h, using on-line ultrafiltration to periodicallyprovide fresh medium. While this process is expensive and may beimpractical for ultra large scales required in the production ofbiofuels, it did demonstrate the possibility of enhancing overallproductivity through supplemental lighting.

Hu, et. al. (1998, Ultra-cell-density culture of marine green algachlorococcum littorale in flat-plate photobioreactor. AppliedMicrobiology Biotechnology: 49, p 655-662) reported producing 84 g/Lalgae in a flat plate photobioreactor with a light intensity of 2,000microeinsteins per second per meter squared. In a review of theliterature up to that time 3-12 g/L were the more typical cell densitiesobtained in photobioreactors.

Fernandez, et. al. (2001, Airlift-driven external-loop tubularphotobioreactors for outdoor production of microalgae: assessment ofdesign and performance Chemical Engineering Science 56 (2001) 2721-2732)reported the results of the design of an airlift system whichincorporated 80 m of clear tubing (pipe) as a solar receiver. The systemwas able to maintain high density algal cultures and used littleexternal power to drive the system. This system's overall design andoperation makes it an attractive candidate for large scale production,but the solar receiver system has limitations for massive scaleproduction.

All current photobioreactor systems have various limitations for use inthe massive scale industrial production of low cost biofuels. Mostsystems are not feasible because of their need for significant amountsof external energy for optimal operation. Another problem with existingsystems is the high cost of the materials to build the systems. Many aremounted on expensive metal racking systems and are not placed on theground. The most significant problem of all existing photobioreactors isthat they are designed to maximize the production of biomass. Theexisting photobioreactors concentrate on the cells reproducing bydivision and increasing in number and mass with the goal of harvestingthe algae or biomass and extracting products form the biomass inseparate steps, or using the biomass itself, or drying the biomass.Current photobioreactor systems have various limitations for use inlarge scale industrial production of low cost biofuels using geneticallyenhanced photosynthetic microorganisms that make biofuels. Most systemsare not feasible because of their need for significant amounts ofexternal energy for optimal operation. Another problem with existingsystems is the complexity and high cost of the materials to build thesystems. Tube or pipe systems are traditionally made from acrylic orpolycarbonate, and that is very expensive and not suited to large scaleproduction. In additional the pipe or tube systems are placed on heavymetal racking to support the liquid culture off the ground, and that isalso very expensive. Existing photobioreactors are designed to maximizethe production of biomass. Existing photobioreactors concentrate on theoptimization of biomass production through reproducing the cells bydivision and increasing in number and mass, or a particular lipid orprotein with the goal of harvesting the microorganisms or biomass fromthe photobioreactor and then extracting the desired products from thebiomass. Harvesting the product from the biomass requires significantenergy and effort and typically requires that the microorganisms orbiomass be replaced for the next cycle of biofuel production.Preferably, a photobioreactor system using genetically enhancedphotoautotrophic organisms for the production of ethanol would collectthe produced biofuel without having to remove the microorganisms orbiomass from the photobioreactor. Traditional photobioreactors areunable to do this as they are designed to produce biomass and this wouldrequire the biomass to be harvested or separated from the water,saccrified and processed in many steps to get and end product of ethanolor lipid. In addition, cell division requires a significant amount ofthe cell's available biochemical energy. It is therefore desirable tomaintain the culture in a steady state and convert as much cellularbiochemical energy to ethanol as possible. Current bioreactor systemsare not typically designed to maintain steady state cultures. Currentindustrial photobioreactor systems do not have a means for trappingethanol produced by genetically enhanced photosynthetic microorganismswhich is released directly into the culture medium. Therefore, incurrent bioreactor systems, the ethanol in the culture medium would bevented to the atmosphere while removing the saturated or supersaturatedlevels of O₂ produced during photosynthesis volatilizing out of theculture medium.

To solve the problem of the inefficiencies involved in recovery ofbiofuels such as ethanol from harvested biomass in photobioreactors, thepresent invention overcomes the external energy usage and materialsconstraints of existing photobioreactor systems while maintaining theneed for culture control and gas exchange necessary for maintaininghigh-density cultures for low cost biofuels production. Current systemshave large requirements for energy to grow and maintain biomass. Thepresent invention uses solar energy and only small amounts of manmadeenergy. The present invention provides for recovery of the biofuel fromcondensate in an upper portion of the photobioreactor and/or from a gasexhaust stream from an upper part of the chamber. The present inventionuses a large airhead space above the culture medium as a means ofallowing the ethanol to evaporate from the culture medium and enter thegas phase of the apparatus. The ethanol can then condense on the wallsof the upper part of the apparatus and run into collection troughs, orthe gas phase can be collected as it leaves apparatus and the ethanolenriched gases can go through external means of collecting the ethanolfrom the gas phase. As O₂ is produced by the organisms in the culture,and any small trace amounts of excess CO₂ introduced into the cultureescape, and as water is changed from a liquid phase to a gas phase, O₂and ethanol gas, in addition to the ethanol in the condensate, will bepushed from the photobioreactor and the ethanol must be recovered inorder to maximize the efficiency of the system. In the present inventionthis can be accomplished predominately with solar energy.

Furthermore, the present disclosure provides for recovery of the biofuelfrom condensate in an upper portion of the photobioreactor and/or from agas exhaust stream from an upper part of the chamber. The apparatus andmethods disclosed herein allow for at least approximately 80% to about100% of the total biofuel product to be recovered by condensation fromthe upper portion of the photobioreactor and/or an exhaust gas streamwith no recovery or at most only about 1% to about 20% of total biofuelproduction to be recovered from the biomass in a lower portion of thephotobioreactor.

The production of biofuels using genetically enhanced photoautotrophicorganisms on an industrial scale requires photobioreactor systems whichcover hundreds, if not thousands, of hectares of land and containmillions, if not billions of gallons of water for organism growth. Suchsystems have never before been developed and pose significantengineering challenges. Because the product being produced is a low costcommodity, not a high value product, low manufacturing costs arecritical for overall commercial success. This one constraint aloneeffectively eliminates all current designs of industrial sizedphotobioreactor systems for the manufacture of biofuels on a largescale. The current invention allows for photobioreactor systems whichcan efficiently produce biofuels from genetically enhancedphotosynthetic microorganisms on a massive scale at low cost.

When exposed to sufficient light, such as sunlight, the geneticallyenhanced organisms release the biofuel into the aqueous growth mediumwhere it evaporates as a gas into the upper part of the chamber. Thewater and biofuel condense on the inner surface of the upper part of thechamber and the droplets run down the internal surface into a collectiontrough, which in one embodiment uses gravity to drain to a lower areafor distillation. Although this evaporation and condensation will happencontinuously during the day, the greatest production of evaporation andcondensation will most likely be at night during the time when there isa greater temperature differential between the inside of the bioreactorand the outside ambient air temperature.

Because the genetically enhanced organisms release the biofuel into thesurrounding growth medium where it evaporates, none or very little ofthe culture has to be harvested or removed to recover the biofuel. Thisresults in significantly increased efficiency and net energy gain fromthe system compared to photobioreactor systems that have to expendresources to remove most or all of the culture from the photobioreactor,separate the biomass from the culture, process the biomass bycentrifugation or saccrification to extract a product, then neworganisms have to be cultured and inoculated, and then the organism isreplaced in the culture in the photobioreactor. None of these steps aretrivial or inexpensive.

The photobioreactors and methods disclosed herein are especiallyapplicable for use as large-scale outdoor photobioreactors, wheresunlight is utilized as the light source. The term “large-scale” inreference to photobioreactors means photobioreactors having a volumegreater than about 1,000 liters, or in some embodiments, greater thanabout 10,000 liters. The photobioreactors comprise closed shapes thatcan be any shape including but not limited to those with rectangular,triangular, cylindrical, circular, oval, irregular or polygonalcross-sections. The photobioreactors can be tube shaped, hexagonal ormultisided domes, or circular domes. The photobioreactor is closed tothe surrounding environment in the sense that loss by evaporation ofbiofuel is kept low and order to prevent: and to prevent contaminationfrom heterotrophic bacteria and other organisms and their waste;evaporation of water; reduction in salinity changes; containment of theorganisms; theft; and vandalism is minimized.

Biofuels able to be produced and released in the present inventioninclude, but are not limited to, ethanol, butanol, pentanol and otherhigher alcohols. In a preferred embodiment, the biofuel is ethanol.

Constructs and methods for producing ethanol from genetically modifiedcyanobacteria have been disclosed (Ref: Woods et al., Fu/Dexter, Colemanet al.). These methods provide for the production and release of liquidethanol into a culture medium from cyanobacteria exposed to sunlight andprovided water, CO₂ and nutrients. Methods and devices exist for thegrowth, maintenance, and harvest of algal or cyanobacterial biomass fromaqueous cultures on a small scale. Most are designed for use in alaboratory and use artificial light to stimulate photosynthesis. Methodsfor separating ethanol from aqueous solutions are also disclosed. Theseseparation devices all require energy in the form of manmade externallygenerated power to drive the separation process, whether it results fromheating (distillation) or cooling (pervaporation). There are no devicescurrently available or described that provide for the large scale,outdoor growth and maintenance of cyanobacterial cultures producingethanol that also enable the separation of ethanol from the aqueousculture in the same apparatus where the cyanobacteria are beingcultured, wherein certain embodiments are made cheaply, are driven bysolar power, use diurnal variation in light and temperature, and aredesigned for controlling the amount of nutrients, light, water, and CO₂to which the cyanobacteria are exposed.

The present invention solves these problems. It provides for an outdoorlarge volume closed photobioreactor for the continued daily productionof ethanol from a culture media comprising genetically enhancedcyanobacteria or algae and separation of the ethanol from the culturemedia through evaporation by sunlight and subsequent condensation andethanol collection in the same photobioreactor chamber in which thecyanobacteria grow. An embodiment of the invention includes the use of asingle chamber composed of translucent plastic which contains a spacefor growing and maintaining the cyanobacteria in an aqueous culturemedium, an airhead for evaporating the ethanol from the aqueous solutionusing sunlight, and a surface for condensing the ethanol from theevaporated gas. This embodiment has troughs for the collection of thecondensed liquid ethanol that can then be moved through attached pipesto a separate apparatus for processing the ethanol to the desiredpurity. In another embodiment the upper part of the chamber of thephotobioreactor has additional coolant compartments in thermal contactwith the gas in the upper part of the chamber. A fluid coolant is passedthrough the coolant compartment that further cools the gas in the upperpart of the chamber and enhances condensation of the biofuel into thecollection troughs. The photobioreactor may be constructed as a singlepiece or as multiple pieces, such as a separate upper part of thechamber and lower part of the chamber, joined together, and in anembodiment is made from lightweight and inexpensive materials, includingrigid materials such as extruded plastic, molded plastic domes, glass,fiber glass, plastic sheets or panels, and flexible materials, such asplastic film, or a combination of flexible and rigid materials. Theupper part of the chamber is optionally coated with a material orconstructed from materials that selectively filter out wavelengths oflight. For example, the upper part of the chamber can be coated orconstructed from a material that filters out potentially harmful UVlight and/or only transmits a specified wavelength range optimal forphotosynthesis by the organisms in the bioreactor.

The photobioreactor can contain ports for the injection of carbondioxide or other gases. The injection of gas is designed to producechurning of the aqueous culture. Churning and mixing in the growthmedium allows higher density cultures and higher biofuel production byminimizing the effects of mutual shading. Churning and mixing alsoprovides for increased gas exchange from the growth medium to the gasphase in the upper part of the chamber and from the gas phase to thegrowth medium. Since oxygen is known to inhibit photosynthesis, removalof the oxygen produced during photosynthesis from the growth mediumhelps to optimize biofuel production. Churning also helps the carbondioxide in the gas phase pass to the growth medium to support carbonfixation and increase biofuel production. Churning can be controlledthrough the use of baffles and dams, mixing devices, injection of gasessuch as carbon dioxide through the growth medium, as well as by theliquid flow through the photobioreactor. Excess oxygen in the growthmedium or in the gas immediately above the culture can inhibit thecellular production of ethanol or other biofuels. Accordingly, ports oroutlets can remove excess oxygen.

Thus, the current invention is a device that provides for the largescale, outdoor growth and maintenance of cyanobacterial culturesproducing ethanol that also enable the separation of ethanol from theaqueous culture in the same apparatus where the cyanobacteria are beingcultured, wherein certain embodiments are made cheaply, are driven bysolar power, use diurnal variation in light and temperature, and aredesigned for controlling the amount of nutrients, light, water, and CO₂to which the cyanobacteria are exposed. One notes that the approachdisclosed herein is not mentioned in the review article by C. U. Ugwu,et al., Photobioreactors for Mass Cultivation of Algae, 99 BioresourceTechnology 4021 (2008).

The above discussion includes both information known to the art prior tothe filing date and information forming part of the present inventivedisclosure. Inclusion of any statement in this section, whether as acharacterization of a published reference or in a discussion oftechnical problems and their solutions, is not to be taken as anadmission that such statement is prior art.

SUMMARY OF THE INVENTION

This invention is directed to a photobioreactor closed from the outsideenvironment comprising a chamber which comprises: a headspace; an upperpart of the chamber which comprises a translucent or clear region toallow in sunlight: a lower part of the chamber which comprises anaqueous growth medium comprising a culture of genetically enhancedorganisms disposed in the growth medium, wherein said organisms areselected from the group consisting of algae and cyanobacteria, andwherein said organisms produce ethanol on a continued daily basis whichenters the growth medium; wherein the ethanol in the growth mediumevaporates into the headspace, condenses on the inner surface of theupper part of the chamber, and collects in a collection trough and thechamber possesses a plurality of openings for inlet and outlet tubes.

An embodiment of the invention is based on the discovery that theethanol concentration in the collection trough can be greater than theethanol concentration within the aqueous growth medium. The presentinvention is distinct from the prior art in that the inside surface ofthe upper part of the chamber is functioning to condense ethanol fromthe gas phase to the liquid phase. Previous approaches, such as thatembodied in Fu and Dexter WO 2007/084477, condense gas phase material toliquid phase material, using only a condenser located outside thechamber. The present invention thus gives a new function to the upperpart of the chamber, a surface upon which gas phase material cancondense to the liquid phase, in addition to the function of allowinglight to enter the chamber. Separately, the result that the condensedethanol in the upper chamber is enriched in ethanol relative to thecontent of ethanol in the liquid phase of the aqueous growth medium ofthe lower chamber is unpredictable and unexpected. Within the chamber,in the limit of obtaining thermodynamic equilibrium, the concentrationsof ethanol would be the same in both liquid phases (that of thecollection trough and that of aqueous growth medium). The collectiontrough of the present invention allows one to capture the unexpectedbenefit.

A further embodiment is an apparatus for the continued daily productionof ethanol and oxygen from carbon dioxide and water in a closed reactionvolume, wherein: the reaction volume comprises a liquid-phase volume anda gas-phase volume; the carbon dioxide and water is converted to ethanoland oxygen in the liquid-phase volume by genetically-modifiedmicroorganisms selected from the group consisting of algae,cyanobacteria; the ethanol, water and oxygen enter the gas phase andoccupy the gas-phase volume; the ethanol and water in the gas-phasevolume are condensed to a condensate volume wherein the concentration ofthe ethanol in the condensate volume is higher than the concentration ofethanol in the liquid-phase containing the microorganisms

A further embodiment is an apparatus for the production of ethanol andoxygen from carbon dioxide and water comprising modular plasticextrusions, at least partially clear or translucent on top, mounted onsoil; a liquid phase reaction volume; a head space volume; a means ofintroducing carbon dioxide to the liquid phase reaction volume; a meansof converting the carbon dioxide into ethanol and oxygen within theliquid phase reaction volume, wherein oxygen goes from the liquid phasereaction volume into the head space volume; a means of separatingethanol from the liquid phase reaction volume by forming gas-phaseethanol followed by condensation of the gas-phase ethanol toliquid-phase ethanol which the condensed liquid-phase ethanol flows to acollection volume.

A further embodiment of the invention is an apparatus with a chambercomprising a container comprising a mixture comprising liquid phaseethanol and liquid phase water, comprising a head space comprising gasphase ethanol and gas phase water, comprising an inner surface on whichgas phase ethanol and gas phase water can condense, and a collectiontrough for collecting said condensed ethanol and water, wherein theconcentration of ethanol in the collection trough is higher than in thecontainer which comprises the liquid phase culture medium.

A further embodiment is a method of producing ethanol comprising:placing a culture of genetically enhanced organisms capable of producingethanol selected from the group consisting of algae, cyanobacteria in aphotobioreactor, wherein said photobioreactor comprises: a) a lower partof the chamber containing an aqueous growth medium, and b) a gas-filledupper part of the chamber, wherein the upper part of the chamber is atleast partially translucent; allowing ethanol to evaporate from growthmedium into the upper part of the chamber; condensing the evaporatedethanol; and collecting the condensed ethanol in one or more collectiontroughs.

BRIEF DESCRIPTION OF THE DRAWINGS

The apparatuses and methods hereof will hereinafter be described inconjunction with the appended drawings, where like designations denotelike elements.

FIG. 1 shows a photobioreactor of the present invention having arectangular tube shape, an upper and lower part of the chamber.

FIG. 2 shows a photobioreactor of the present invention having ahexagonal dome shape.

FIGS. 3A and 3B shows alternative photobioreactor designs for a tubeshaped photobioreactor similar to the photobioreactor of FIG. 1. Thealternative designs depicted in FIG. 3 do not contain means for heatexchange below the growth medium.

FIG. 4A shows a photobioreactor having two upper coolant compartmentsextending across the upper part of the chamber from the sides. FIG. 4Bshows a photobioreactor where the upper coolant compartment is a flatpanel along the side of the chamber.

FIGS. 5A and 5B shows alternative tube designs of the photobioreactorsimilar to the photobioreactor of FIG. 1.

FIGS. 6 and 7 also show three-dimensional views of different shapes anddesigns of photobioreactor tubes suitable with the present invention.

FIG. 8 shows a three-dimensional view of the ends of twophotobioreactors placed adjacent to one another.

FIG. 9 shows hexagonal dome shaped photobioreactors fused togetherhaving multiple positions for gas and nutrient inlets and outlets.

FIG. 10 shows a rear, cross-sectional and front view of end plugs usedto seal the inlets and outlets of a photobioreactor similar to thanshown in FIG. 1.

FIG. 11 shows a three-dimensional view of the end plug of FIG. 10installed at the end of a tube shaped photobioreactor.

FIG. 12 shows a three-dimensional view of a photobioreactor flowconnector unit that is used to join separate tube shapedphotobioreactors.

FIG. 13 shows a front and rear view of the flow connector unit of FIG.12.

FIG. 14 shows three-dimensional, end and cross-sectional view of aphotobioreactor coupling unit that is used to join two separate tubeshaped photobioreactors with the flow continuing in one direction.

FIG. 15 shows a photobioreactor coupling unit installed on onephotobioreactor ready to receive a second photobioreactor.

FIGS. 16-19 show the top and side views of various designs andconfigurations of flow dams and baffles placed at the bottom of aphotobioreactor utilizing a pump or flow of liquid through the lowerchamber.

FIG. 20 shows a three-dimensional view of various tube fittings used toconnect the inlet and outlet tubes for photobioreactors of the presentinvention.

FIGS. 22A and 22B illustrate the gas exchange above and below the growthmedium and different phases of ethanol in a photobioreactor. FIGS. 21Aand 21B illustrate the turbulence and gas exchange in twophotobioreactors having baffles and dams.

FIG. 23 illustrates a photobioreactor made of tubular polyethylenematerial with condensate collection troughs manufactured into the finalphotobioreactor.

FIG. 24 illustrates various means to manufacturing condensate collectiontroughs into said photobioreactor.

DETAILED DESCRIPTION

Disclosed herein are photobioreactor systems and methods that permit theculture of genetically-enhanced photoautotrophic organisms that canachieve high biomass densities and high photosynthetic conversion rates.In addition, combinations of known technologies that can controltemperature, nutrients, gases and waste can be used herein. Such aclosed, large-scale, large-volume, external solar receiver apparatus foroutdoor photobioreactor systems for industrial scale production has notbeen shown heretofore, using natural sunlight to drive photosynthesisfor biomass production and requiring minimal amounts of external energyto operate.

The design and operation of the total photobioreactor system isoptimized for the removal of excess heat, to maintain correct pH,salinity, maximum ethanol evaporation, minimal water evaporation, toreduce or eliminate water loss to the environment, to limit thepossibility of contamination and to permit the development of asubstantial concentration of a biofuel such as ethanol in the growthmedium, and to limit spontaneous mutations in the cultured organisms,and to regulate temperature, to maximize the total amount of organismsin the growth medium, to maximize the total amount of organisms in thegrowth medium that receive light by churning the organisms in the growthmedium from the top to the middle and bottom and back to the top, tomaximize the photosynthetic rate of the organisms by limiting lightsaturation and minimizing dark or shaded periods in the tube, and tobalance the effect of saturation and shading, and to limit the effect ofspontaneous mutations in the cultured organisms.

An embodiment of the photobioreactor provided herein has a shape anddesign that maximizes the incidence of solar energy absorption duringphotosynthesis. The photobioreactor design is engineered to reduce thedaily variation of the temperature when placed on location at theproduction facility.

Designs for photobioreactors in the prior art are intended for theproduction primarily of biomass, and secondarily of products derivedfrom the biomass. They require inputs of carbon dioxide, water, mineralnutrients and light. They have outputs of oxygen and biomass. As apractical matter they may also have outputs into the growth medium,which must be separated from the biomass, water vapor, and incidentalcomponents of the gas phase such as nitrogen. Methods have beendisclosed for the production of hydrogen using cultures of organisms ina photobioreactor designed and operated for that purpose. Hydrogenproducing photobioreactors provide an anaerobic environment in whichhydrogen production occurs. There may be internal production of biomassthat is consumed as part of the hydrogen production process. The onlyessential output is hydrogen, and the only essential inputs are waterand light. The ethanol producing photobioreactor system of the presentinvention has inputs of carbon dioxide and water and light. As apractical matter, it may be necessary to introduce some mineralnutrients into the reactor to allow enough growth for cell replacement,but addition of mineral nutrients is not inherent in the design of theprocess as it is in a biomass producing photobioreactor. A method ofreducing heterotrophs can be accomplished with a nutrient deficientgrowth medium.

The present invention provides for an outdoor, large volume, closedphotobioreactor for the continued daily in situ production of ethanolfrom genetically enhanced cyanobacteria, or algae in culture, release ofethanol by evaporation from the culture into the headspace of thedevice, collection of ethanol by condensation, and removal of theethanol through collection troughs in the device. Specifically, thephotobioreactor comprises a chamber which comprises an upper part of thechamber which comprises a translucent or clear region to allow insunlight and an inner surface of the upper part of the chamber on whichinner surface ethanol and water may condense. The photobioreactorcomprises a lower part of the chamber which comprises an aqueous growthmedium comprising a culture of genetically enhanced organisms disposedin the growth medium. The chamber comprises a plurality of collectiontroughs for the collection of liquid phase materials. The ethanolcollected in the collection trough can be of higher concentration thanthe concentration of ethanol in the growth medium.

The photobioreactor may be made of clear plastic or other translucentmaterial molded into desired shapes such as extruded tubes or moldeddomes. The top of the photobioreactor is clear to allow sunlight throughand photosynthesis to occur in the cyanobacteria or algae in culture.The enhanced cyanobacteria or algae in culture produce the ethanolintracellularly from sunlight, CO₂, water, and then release it to theculture medium. The photobioreactor is designed to promote evaporationof ethanol from the culture medium and then condense the ethanol into aliquid that can be captured in troughs and removed for furtherdistillation. The upper inside part of the photobioreactor is where theethanol condenses and collects to run to the troughs. The upper part ofthe chamber of the photobioreactor may have additional internal coolantchambers with fluid coolant passing through them to maximize thecondensation of ethanol for collection in the troughs. Thephotobioreactor has inlets that allow for the introduction of water,CO₂, and nutrients, and outlets for the removal of ethanol, water,by-products, and O₂. The photobioreactor could be made without theinternal coolant chambers including when conditions do not require thelower temperature to condense the ethanol. The upper part of the chambermay be divided into sections to allow heat to rise above and away fromthe culture. The upper part of the chamber may have materials applied tothe surface that maximize the condensation of ethanol. Thephotobioreactor may be made in one piece or in multiple parts andassembled.

The lower part of the chamber contains the genetically modifiedcyanobacteria, algae or other photoautotroph culture that produces theethanol. The lower part of the chamber containing the culture is shapedto allow for the easy mixing of the culture and nutrients by thebubbling of CO₂ and the increased dissolving of the CO₂ gas into theculture. The lower part of the chamber is also shaped to allow for themaximizing of the surface area and therefore contact with the bottomcompartment filled with temperature regulating fluid and would serve tocool the culture in the heat of the day, and keep the culture warm atnight. The lower part of the chamber also has rails to allow for the useof a mixing device if necessary. The lower part of the chamber hasinlets incorporated in the sides and ends to allow for the introductionof nutrients and the return of water. The lower part of the chamber hasmultiple inlets incorporated in the bottom to allow for the bubbling ofCO₂ gas. The bottom chamber, when needed, is filled with temperatureregulating fluid. The photobioreactor could be made without thetemperature maintenance compartment under the lower part of the chamberfor use in moderate climates, but with the temperature maintenancecompartments and the temperature regulating fluid, the photobioreactorcan be adapted to higher temperatures in many marginal land and desertareas. The regulating properties of the photobioreactor allow for theproper conditions to be regulated and maintained for maximizing ethanolproduction during different times of the day and during changingseasons. In addition, the photobioreactor design permits the regulationof internal nutrients, by-products, gases, and optimizing ethanolproduction conditions in the culture. The photobioreactor can haveculture monitoring and regulating devices attached to it.

The liquid growth medium can be any growth medium useful for themaintenance of a culture of cyanobacteria, and algae. For example, itcan be an aqueous growth medium such as water only, water and carbondioxide, or a solution comprising water, carbon dioxide and nutrientsthat are helpful or necessary for maintaining the culture. Thephotobioreactor has inlets that allow for the introduction of water andnutrients, and culture monitoring, measuring devices and regulatingdevices that can detect when the water and/or nutrient level in thelower part of the chamber fall outside a certain predetermined level. Acontroller takes corrective action and activates the pump or valve toregulate pumping or passing the water and/or nutrients through the inletand into the lower part of the chamber to keep the levels within acertain predetermined range.

The current invention eliminates the need for such gas exchange systemsby using a significant airhead (gas phase) space contacting the physicalculture. When combined with a low energy mixing system and/or means forintroduction of CO₂ by a bubbling system, CO₂ can be injected for use bythe organisms and the O₂ released from the liquid phase to the gas phasespace.

The design of the photobioreactor apparatus as part of a totalbioreactor system is optimized for the removal of excess heat, tomaintain correct pH and temperature, to limit evaporation to theexternal environment, to limit or control changes in salinity, to limitthe possibility of contamination an and to permit the development of asubstantial concentration of a biofuel such as ethanol in the growthmedium, and to limit spontaneous mutations in the cultured organisms.The photobioreactor apparatus has a sufficient size to maximize solarenergy absorption by the culture to facilitate maximum photosynthesiswhile covering the ground surface in an efficient manner to maximizephotobioreactor apparatus surface area, maximize total facility groundcoverage, minimize supply and service areas to lower overallinstallation cost.

The photobioreactor apparatus comprises an upper portion composed of atransparent or translucent material to allow for the passage of light tothe inside of the apparatus. The inner face of the upper portion will bein contact with the gases supplied to the culture system, as well as thegases produced during photosynthesis. The lower portion of thephotobioreactor apparatus may be made from the same material as theupper portion, but does not have to be composed of a transparent ortranslucent material.

During operation of the photobioreactor, cells can adhere to the wallsof vessels and exhibit foaming. Foaming creates air pockets and celladherence to the photobioreactor walls restricts solar light penetrationand, thus, uniform absorption by the cells. Foaming may be mitigatedthrough the use of antifoam agents, such as polypropylene glycol (PPG).Additionally, the inner surface of the photobioreactor is optionallytreated with an anti-fouling material to prevent cell adherence.

Where ethanol production is best optimized by the mixing of the culturethen several options exist in this photobioreactor, a mixing device maybe added, or the culture may be pumped into one end where the apparatuscan further comprise internal mixing means. For example, static mixerscomprise of a series of internal flow baffles, which mimics the mixingand churning in a stream bed, while minimizing shear damage to thecultured organisms, to provide turbulent flow and appropriate mixing andchurning of the culture to ensure uniform exposure of all of the cellsto solar radiation. Moreover, the turbulent flow thus produced not onlyprovides mixing for maximal photosynthesis, but also serves to removedissolved oxygen and enhances absorption of carbon dioxide from theairhead above the culture. Internal static mixers or baffles can be usedon systems which are installed on properly sloped surfaces that providesufficient liquid flow to achieve the desired mixing and churning aswell as sufficient residence time in the system to cost effectivelyproduce ethanol. Static mixers are appropriate if a means of circulationis provided so that a flow of water exists. Static methods are onlyeffective if there is a water flow. Optionally, the photobioreactorapparatus may be fitted with dams to control the flow of culture throughthe apparatus in areas where there is a slope to the surface where thephotobioreactor is situated.

In physical locations where the land is flat or only slightly sloped, orif recirculation pumps are not used, it may be necessary with certaingenetically enhanced photoautotrophic cyanobacteria or algae, to installan internal mixing systems. Such systems can have numerous designs, butthey must be able to operate using minimal external energy whileproviding sufficient mixing for optimal ethanol production and gasremoval. There are a wide variety of means for mixing fluids which couldbe adapted to mixing the culture within the reactor well known to thoseskilled in the art. For example those mixing means used in the design ofnon-photosynthetic bioreactors and operated at energy levels appropriatefor fuel production could be used. Other mixing means not now used inbioreactors may also be applicable. The correct use of such mixerseliminates the high energy consumption associated with pumpedcirculation of growth medium common to many bioreactor designs.

But in most instances with genetically enhanced photoautotrophiccyanobacteria or algae, the photobioreactor is designed to maintain astatic culture at high cell density, with minimum cell growth anddivision, with much of the energy flow from photosynthesis going toethanol production and not mere biomass accumulation.

Excess oxygen must be allowed to escape. This can be accomplished byventing the gas phase. A small amount of ethanol will also be lost whenoxygen is vented, unless oxygen venting uses one or more of the ethanolretention methods described below. In the event oxygen is particularlyinhibitory to the organism in culture, it may be desirable to furtherreduce the oxygen level in the photosynthetic enclosure. This may beaccomplished by several means. For example, a carrier gas such as normalair can be passed through the system so that the venting is more rapidthan it would be by build up of oxygen pressure alone, so the steadystate oxygen concentration is between normal atmospheric levels and 300%oxygen, with the actual level determined by the system design.Additionally the entire gas phase may be removed mechanically from abovethe culture and passed through a number of existing devices that extractethanol from the gas phase, including a dephlegmator, cooling heatexchangers, or other forced extraction methods from the gas phaseoutside of the photobioreactor. Non-venting methods are also available,such as passing the gas phase past an oxygen permeable membrane. Reducedpressure, e.g. provided by a vacuum pump on the other side of themembrane provides a pressure gradient to drive oxygen, but not other gasphase components, through the membrane.

Since genetically enhanced photoautotrophic aquatic organisms releasethe ethanol they produce directly into the growth medium, there is asignificant technical challenge to extract the ethanol using minimalexternal energy. One way to harvest the ethanol is to take some of thegrowth medium, and use well-established industrial ethanol purificationmethods, (distillation, for example) resulting in the production ofethanol, a waste water stream, and a dried residue that may have valuein feed or other applications. If growth medium is removed, it must bereplaced with fresh medium. Additional algae should be added to make upfor that which was removed, unless internal growth is sufficient to makeup for removed algae. Harvesting biomass from which to recover thebiofuel requires external energy for moving the growth medium/biomassmixture, for physical organism separation, and separate biofuelextraction. The cost and energy required for this type of process makesit unfeasible for large-scale use. The present disclosure provides analternate means to remove the biofuel from the photobioreactor withoutthe need for moving the growth medium, pumping separating the organismsby filtration or centrifugation, producing the biofuel by saccrificationand further processing. Although this is process can be done, it willrequire a lot of external energy for moving the growth medium/biomassmixture, for physical organism separation, and separate ethanolextraction.

The invention provides for an alternate means to remove the ethanol fromthe growth medium without the need for pumping and/or filtration usingonly the energy from the sun. This is accomplished by providing for asignificant airhead (gas phase) space above the culture. This spaceallows the sun to cause evaporation of the ethanol and water into theairhead space. Since the space inside the bioreactor is hotter than theoutside ambient air temperature, the water and ethanol vapor willcondense on the inner surface of the top surface of the bioreactor. Thewater and ethanol vapor condensates on the inner surface of thephotobioreactor apparatus and the droplets run down the internal surfaceand fall into a collection trough, which uses gravity to drain to acentral collection area for distillation. Although this evaporation andcondensation will happen all day, the greatest production will be atnight during the time when there is a greater temperature differentialbetween the inside of the bioreactor and the outside ambient airtemperature. This method of removing the ethanol from the culturewithout moving or disturbing the overall culture is very important inreducing the overall energy needed for the production system.

If there is excess condensate, the excess can be returned to thebioreactors either as a fluid or evaporated in a gas stream. Anotheralternate method of ethanol harvest is to pump the gas phase to anexternal condensation unit that may serve one or many bioreactors, andprovide a cooled surface on which condensation may occur, subsequentlyreturning the cooled and ethanol depleted gas phase to the bioreactors.Cooling may be provided by any economical means, and energy costs may beminimized if desired by use of heat exchangers or the like. Thetemperature to which the gas stream is cooled must be less than thetemperature of the bioreactors to get condensation, but it need not beso low that all or a large fraction of the ethanol is removed from thegas stream, since the gas stream in this case is not vented to theatmosphere, ethanol that is not condensed to liquid is not lost butreturned to the system. The condensate collected in this manner wouldthen be further purified by distillation or other means. A furthermodification of this method of ethanol collection and removal which maybe valuable would be to pass the gas phase or a portion of the gas phasethrough a cold trap, which would result in the removal of essentiallyall of the ethanol from the gas phase. This portion of the gas phasecould then be vented to the air to allow for removal of excess oxygen,or the oxygen could be collected and used. Heat exchangers and the likecan be used to reduce energy costs associated with cooling. It may beworthwhile in the process of harvesting ethanol using the above methodsto use a device such as a dephlegmator, which passes the gas phasethrough a temperature gradient with a condensation and evaporationprocess reminiscent of a still which results in the production of aliquid stream enriched in water and a gas stream enriched in ethanol, sothat the condensed ethanol has greater purity and subsequentpurification is simpler and less costly. It may prove most economical touse a combined approach, in which most ethanol is removed using thephotobioreactor internal condensation, with a relatively highcondensation temperature and very low operating costs for the bulk ofethanol removal, and to use the low temperature condenser with optionaldephlegmator to process the smaller gas volume that must be removed toget rid of excess oxygen. It can be worthwhile in the process ofharvesting ethanol using the above methods to use a device such as adephlegmator. Such dephlegmator are known to the art, and pass the gasphase through a temperature gradient with a condensation and evaporationprocess reminiscent of a still, which results in the production of aliquid stream enriched in water and a gas stream enriched in ethanol, sothat the condensed ethanol has greater purity and subsequentpurification is simpler and less costly. Those skilled in the art willbe able to design an optimal system using these components that providesfor ethanol harvest and oxygen removal.

In geographic locations where internal heat in the photobioreactorapparatus has a negative affect on the culture growth, it may benecessary to have a separate compartment below the culture in theapparatus to have water that can be used to cool or heat the overallculture. This extra mass of water can also be used to regulate thenatural day to night heat and cooling cycle in locations with wide dailytemperature swings such as desert locations. Additional cooling may benecessary, especially during periods of hot weather or the midday highsun. Cooling can be provided by water evaporation, for example byplacing or submerging the photobioreactor apparatus in a pond containingwater, said pond having a surface exposed to the air so that evaporativecooling occurs. The photobioreactor apparatus is in turn cooled bycontact with the cooling water. Many different arrangements arepossible. Photosynthetic enclosures may be floated on the water surface,submerged or partially submerged, movable within the pond, or the waterdepth of the pond may be adjusted to get any desired degree of heattransfer and evaporative cooling. Rates of heat transfer, evaporation,pond temperature as a function of weather conditions and the like can bepredicted by those skilled in the art and used to obtain the desiredtemperature in the photosynthetic enclosure. Alternatively, coolant,water or other convenient material, that can be passed through thebottom temperature maintenance compartment, may be pumped, flowed, orpassed to a processing site and cooled by other means such as anevaporative cooling tower. Alternatively, water may be pumped from acool location such as a deep ocean location, used for cooling, and thendiscarded.

Furthermore, it is not necessary to continue to increase the biomass ofthe culture in some embodiments. When the culture has reached a suitablesize for the photobioreactor, the cellular activities should shift awayfrom continued replication, which demands greater nutrient presence, andtoward biofuel production.

Enclosures may be made of rigid materials such as extruded plastic,molded plastic domes, or plastic sheets or panels, or flexiblematerials, such as plastic film, or a combination of flexible and rigidmaterials. It may include framing members to impart strength or form tomaterials such as plastic extrusion, panels, or film that wouldotherwise have inadequate mechanical properties to create the desiredstructure.

The bioreactor must be provided with CO₂ and water to provide substratefor the photosynthetic conversion of CO₂ to sugar, which is thensubsequently converted to biofuel within the cells and then dispersedinto the medium. CO₂ can readily be introduced as a gas, either into thegas phase or bubbled into the medium, or by means of a liquid where theCO₂ is supersaturated dissolved in the liquid.

The photobioreactor may be constructed as a single piece or as multiplepieces, such as a separate upper part of the chamber and lower part ofthe chamber, joined together, and in addition, lower temperaturemaintenance compartments may be joined to comprise the apparatus, and inan embodiment nutrient requirements of a biofuel producingphotosynthetic apparatus are preferably low, since the culture should beproducing primarily biofuels devoid of mineral nutrient content.Nutrients may leak out of cells into the medium and be re-used. Unlikeother nutrients, nitrogen is often present in low but significant levelsin volatile form, so there may be a gradual loss of ammonia from theculture in the photosynthetic enclosure. If this is the case, it can bereplaced, either in the gas or liquid phase.

The lower portion of the photobioreactor apparatus can be made from thesame material as the upper portion, but does not have to be composed ofa transparent or translucent material. Additionally, the apparatus canbe made airtight and should be watertight, and all connections andfittings in the system should be kept to a minimum, and be designed toprevent contamination of the culture.

The upper part of the chamber or a significant portion of the upper partof the chamber is transparent or at least partially translucent. As usedherein, “partially translucent” should be understood as permittingsufficient passage of light, particularly sunlight, into thephotobioreactor to enable photosynthesis by photoautotrophic organismswithin the photobioreactor. In embodiments hereof, the upper part of thechamber or a significant portion of the upper part of the chamber isclear, transparent, or partially transparent. The upper part of thechamber is optionally coated with a material or constructed frommaterials that selectively filter out wavelengths of light. For example,the upper part of the chamber can be coated or constructed from amaterial that filters out potentially harmful UV light and/or onlytransmits a specified wavelength range optimal for photosynthesis by theorganisms in the bioreactor. Such materials used in construction andcoatings of transparent and translucent devices are well known in theart.

The photobioreactor apparatus has a sufficient size to maximize solarenergy absorption by the culture to facilitate maximum photosynthesiswhile covering the ground surface in an efficient manner to maximizephotobioreactor apparatus surface area, maximize total facility groundcoverage, and minimize supply and service areas to lower overallinstallation cost.

During operation of the photobioreactor, cells can adhere to the wallsof vessels and exhibit foaming. Foaming creates air pockets and celladherence to the photobioreactor walls that restricts solar lightpenetration and, thus, uniform absorption by the cells. Foaming can bemitigated through the use of antifoam agents, such as polypropyleneglycol (PPG) and other antifoam agents known to the art. Additionally,the inner surface of the photobioreactor is optionally constructed ortreated with an anti-fouling material as known in the art to preventcell adherence.

Also during operation of the photobioreactor, cells can adhere to thewalls of vessels and it is also known in the art that certainhydrophobic and hydrophilic substances promote beading or collection ofcondensing liquids on a surface. The inner surface of the upper part ofthe chamber is optionally constructed or treated with such a substanceto promote condensation in the upper part of the chamber.

Churning and mixing in the growth medium allows higher density culturesand higher biofuel production by minimizing the effects of mutualshading. Churning and mixing also provides for increased gas exchangefrom the growth medium to the gas phase in the upper part of the chamberand from the gas phase to the growth medium. Since oxygen is known toinhibit photosynthesis, removal of the oxygen produced duringphotosynthesis from the growth medium helps to optimize biofuelproduction. Churning also helps the carbon dioxide in the gas phase passto the growth medium to support carbon fixation and increase biofuelproduction. Churning can be controlled through the use of baffles anddams, mixing devices, injection of gases such as carbon dioxide throughthe growth medium, as well as by the liquid flow through thephotobioreactor.

In one embodiment, a photobioreactor comprises an internal mixing meansin addition to the carbon dioxide bubbling means. Providing mixing orchurning in the growth medium ensures uniform exposure of all of thecells to solar radiation and evenly distributes nutrients and CO₂ in thegrowth medium. Moreover, the mixing and churning not only providesmixing for maximal photosynthesis, but also serves to remove dissolvedoxygen from the growth medium and enhances absorption of carbon dioxidefrom the gas phase above the culture. Such churning and mixing systemscan have numerous designs, but they should be able to operate usingminimal external energy while providing sufficient mixing for optimalbiofuel production and gas removal. There are a wide variety of meansfor mixing fluids, such as stirrers, which can be adapted to mixing theculture within the reactor by means well known to those skilled in theart. For example those churning and mixing means used in the design ofnon-photosynthetic bioreactors and operated at energy levels appropriatefor fuel production could be used. Other mixing means used in fieldsother than bioreactors can also be applicable. The correct use of suchmixers eliminates the high energy consumption associated with pumpedcirculation of growth medium common to many bioreactor designs. Thelower part of the chamber can comprise rails or other support structuresto allow for the use of a churning and mixing device. The lower part ofthe chamber is also optionally shaped to allow for the easy churning andmixing of the culture and nutrients by the bubbling of CO₂ and theincreased dissolving of the CO₂ gas into the culture, such as by havinga corrugated, sloped, or peaked bottom surface (as shown in FIGS. 1, 3a, and 4).

In a further embodiment, the photobioreactor can comprise static mixers,such as a series of internal flow baffles, which mimic the mixing andchurning in a stream bed, while minimizing shear damage to the culturedorganisms. Internal static mixers such as baffles can be used in systemsthat are installed on sloped surfaces to provide sufficient liquid flowto achieve the desired mixing and churning as well as sufficientresidence time in the system to cost effectively produce ethanol. Staticmixers are appropriate if a means of mechanical circulation is providedso that a flow of liquid exists on flat lands. Optionally, thephotobioreactor apparatus can be fitted with dams to control the flow ofculture through the apparatus in areas where there is a slope to thesurface where the photobioreactor is situated.

Excess oxygen in the growth medium or in the gas immediately above theculture can inhibit the cellular production of ethanol or otherbiofuels. Accordingly, excess oxygen should be removed. This can beaccomplished by venting or removing the gas from the upper part of thechamber. Enough oxygen should be removed to prevent inhibition ofbiofuel production below levels that are economically sustainable. Forexample, oxygen concentration within the lower part of the chambershould generally be kept below the level that would cause oxygeninhibition in that particular culture. Gas can be removed mechanicallyfrom above the upper part of the chamber culture through an oxygenexhaust outlet positioned in the upper part of the chamber.Additionally, gas, including excess oxygen and evaporated biofuel, canbe removed from the upper part of the chamber and passed through anumber of gas collecting devices known to the art that extract thebiofuel from the gas phase, including a dephlegmator, cooling heatexchangers, or other forced extraction methods from the gas phase, whichcan be located outside of the photobioreactor. Non-venting methods arealso available, such as passing the gas phase past an oxygen-permeablemembrane. Reduced pressure, e.g., provided by a vacuum pump on the otherside of the membrane, provides a pressure gradient to drive oxygen, butnot other gas phase components, through a membrane separator. Excesspressure in the photobioreactor could have the same effect and push O₂from inside the photobioreactor. Additionally, a preferred embodiment ofthe apparatus may be made airtight and must be watertight. In addition,all connections and fittings in the system should be kept to a minimum,and must be designed to prevent contamination of the culture.Additionally, a preferred embodiment of the apparatus is that one ormore inlet or outlet, supplies, and drains tubes, may serve one or morefunctions.

In addition, to provide a large-scale photobioreactor while ensuringthat the energy required to produce the biofuel product is less than theenergy value of the produced biofuel, it is necessary to minimizemanmade energy inputs for the creation of the biofuel, and recovery ofthe biofuel. No current photobioreactor uses solar energy to create,separate and recover the biofuel. The present invention can beaccomplished predominately with solar energy.

In no existing photobioreactor does there exist the capability ofproducing a biofuel that is released and volatilized from the culture,removed and collected from the photobioreactor without harvesting thealgae biomass. The key features that distinguish the invention fromprevious bioreactor designs are: (1) It is intended to produce biofuelsdirectly, with the production of little or no biomass, nor harvest ofthe biomass (2) The biofuels produced, such as ethanol, may be volatile,so provision is made to prevent excess loss of biofuels in the gasstreams associated with the removal of oxygen (3) Costs including energyinputs are reduced so that fuels can be offered at a price substantiallyless than those available from other bioreactor systems (4) Unlike cornand sugarcane derived ethanol that are separately grown, harvested,processed, and then their sugars are converted into ethanol in separateprocesses, the invention allows for a single container to grow theethanol producing genetically enhanced photoautotroph, release theethanol, collect, and remove the ethanol.

The chamber is the structure of the apparatus that contains both theliquid culture media and the headspace. The volume of the headspace isdefined by the contours of the inside of the upper part of the chamberand the upper surface of the liquid culture media, as shown in FIG. 1.Ethanol and H₂O gas contained within the headspace condense on the innersurface of the upper part of the chamber.

The headspace contains the gases above the culture and would includeeach of air, CO₂, O₂, H₂O and ethanol.

The upper part of the chamber contains the headspace and the condensate.The upper part of the chamber can be divided into multiple smallerparts. The upper part of the chamber can contain coolant compartmentsthat further facilitate condensation.

The coolant compartments of the upper part of the chamber are in thermalcontact with the headspace in the upper part of the chamber, andcomprising a coolant, wherein the coolant could be selected from thegroup including water, seawater or other coolant known to a person ofordinary skill in the art.

The lower part of the chamber contains the liquid culture media and thiscomprises the water, seawater, brackish water or polluted water, thegenetically enhanced cyanobacteria or algae, the nutrients which couldbe selected from the group including nitrogen, nitrates, phosphates,ammonium, BG-11, CO₂, carbonates, trace elements, agents to promote thegrowth of the organisms, solutes selected from the group consisting ofnutrients, fertilizers, antibiotics, and algaecides.

A list of measuring devices and sensors shall include pH, CO₂, O₂,temperature, salinity, and ethanol probes and sensors, and others knowto a person skilled in the art.

The reason for using UV blocking and stabilizing agents, coatings orfilms, incorporated into the plastic or on the surface of the plastic isto include reducing heat in the culture, and breakdown of the plastic,and mutations of the organisms.

The design and operation of the total photobioreactor system isoptimized for the removal of excess heat, to maintain correct pH,salinity, evaporation and temperature, to reduce the possibility ofcontamination, to maximize the total amount of organisms in the growthmedium that receive light by rolling the organisms in the growth mediumfrom the top to the middle and bottom and back to the top, to maximizethe photosynthetic rate of the organisms by limiting light saturationand minimizing dark or shaded periods in the pipe, or to balance theeffect of saturation and shading, and to limit the effect of spontaneousmutations in the cultured organisms.

The preferred embodiment of the present invention is a length and designthat maximizes the incidence of solar energy absorption duringphotosynthesis. The photobioreactor geometry is engineered to reduce thedaily variation of the solar irradiation when placed on location at theproduction facility. The headspace over the culture also serves tomoderate the heat during periods of high solar radiation.

As part of the photobioreactor system, internal static mixers provideappropriate mixing, which is critical to ensuring uniform exposure ofcells to solar radiation. Linear flow through solar tubes can reduce theoverall photosynthesis since cells near the middle or bottom of thetubes receive less light than those towards the top. The presentinvention overcomes this limitation through the use of internal flowbaffles designed to facilitate mixing. These flow baffles are static innature, thus requiring no additional energy input beyond the flow ratedetermined by gravity. The careful balancing of mixing in the pipes withcreating turbulent flow is important, since the algae cells aresensitive to sheer damage during heavy mixing. The design of thephotobioreactor apparatus mimics the mixing and churning in a naturalstream lined with rocks. The turbulent flow in the photobioreactor notonly provides the necessary mixing for maximal photosynthesis in highdensity cultures, but also serves to remove dissolved oxygen andenhances absorption of CO₂. This also helps to stabilize the culture pHand increases the resident time of the biomass in the photobioreactorfor photosynthesis.

The photobioreactor may also be fitted with dams of varying heights thatcan be used to control the flow of the culture in areas where there is agreater natural slope. This permits steeper surfaces to be used forproduction facilities and lower overall installation costs.

In addition, the dams can increase churning for cultures which grow inhigher densities and need more mixing. The dams also serves as a controlmechanism for growth medium and photoautotrophic organism output bymaintaining an output at the end of the photobioreactor equal to inputat the beginning of the photobioreactor while on a slope.

The dam serves as an inexpensive control mechanism for the time theorganism spends in the photobioreactor tube. The dams also serve as acontrol of growth medium pressure that would occur at the bottom of aslope in a normal pipe. The dam allows for the photobioreactor to beplaced on a slope and eliminates the need for mechanical mixers andstirrers that add to the cost the biofuels production. The growth mediumand photoautotrophic organism can be pumped or poured by gravity intothe top of the sloped apparatus without further need for mixing, orpumping while in the growth phase.

A proper functioning photobioreactor system should be airtight and watertight. Thus, the preferred embodiment of the invention has the number ofconnections and fittings in the total system kept to a minimumpreferably by extruding the photobioreactor on site in long sections.Preferably, such section may be 10-300 feet in length. Other methods ofmanufacturing may be employed such as molding the tubes.

The preferred embodiment of the invention may be fabricated from anymaterial, including glass, but preferably a plastic that has the opticalclarity to permit photosynthesis and can withstand long-term UVradiation exposure and exposure to corrosive saltwater, heat and cold,and expansion and contraction. Glass, and opaque or translucent plasticsmay also be used, as long they meet the needs of the photoautotrophicorganisms to be grown in the system. Any person skilled in the art ofthermoplastics can specifically design a plastic, or plastic mix whichcan be used for the photobioreactor tube. Virgin resins may be used tomanufacture the tubes, but since cost is a likely significant factor,recycled plastics are preferred. A few of the particular plastics caninclude High Density Polyethylene (HDPE), Polyethylene Terephthalate(PET), acrylic, Lucite, polypropylene and polycarbonate. The location ofthe facility, the desired cost of the facility, the desire life-span ofthe photobioreactor are all factors that must be considered beforedeciding on the type of material to use.

Designs for photobioreactors in the prior art are intended for theproduction primarily of biomass, and secondarily of products derivedfrom the biomass. They require inputs of carbon dioxide, water, mineralnutrients and light. They have outputs of oxygen and biomass. As apractical matter they may also have outputs into the growth medium,which must be separated from the biomass, water vapor, and incidentalcomponents of the gas phase such as nitrogen. Methods have beendisclosed for the production of hydrogen using cultures of organisms ina photobioreactor designed and operated for that purpose. Hydrogenproducing photobioreactors provide an anaerobic environment in whichhydrogen production occurs. There may be internal production of biomassthat is consumed as part of the hydrogen production process. The onlyessential output is hydrogen, and the only essential inputs are waterand light. The ethanol producing photobioreactor system of the presentinvention has inputs of carbon dioxide and water and light. As apractical matter, it may be necessary to introduce some mineralnutrients into the reactor to allow enough growth for cell replacement,but addition of mineral nutrients is not inherent in the design of theprocess as it is in a biomass producing photobioreactor. The presentmethods of producing ethanol and other biofuels can be performed with anutrient deficient growth medium.

Unlike the hydrogen producing photobioreactor, carbon dioxide isrequired. Furthermore, in contrast to a biomass producingphotobioreactor, the ethanol producing photobioreactor of the presentinvention has outputs of oxygen and ethanol and does not require thanthe culture be harvested to recover any produced biofuel. As a practicalmatter, there may be some discharge of biomass from the photobioreactorsof the present invention to prevent the development of cultures withexcessive cell density, or to collect a test sample, but the productionof the biomass is not a desired feature. Unlike the hydrogen producingphotobioreactor, the ethanol producing photobioreactor does not producehydrogen, but produces both oxygen and ethanol. Furthermore, while theethanol producing photobioreactor of the present invention may beoptimized to remove excess oxygen, it does not require anaerobicconditions as required by hydrogen photobioreactors.

Both hydrogen producing photobioreactors and biomass producingbioreactors are tolerant of heterotrophic contaminants that are notdisease organisms. Some methods even call for the addition ofheterotrophs. The hydrogen and biomass products are not susceptible tobiological degradation. In contrast, ethanol is readily degraded by manyheterotrophs in the presence of oxygen, so it is essential that suchheterotrophs be excluded or their growth otherwise prevented in theethanol producing photobioreactor. All liquid and gases entering thephotobioreactors of the present invention can be sterilized using UVand/or filtration to reduce the possibility of contamination by foreign,unwanted microorganisms.

The overall size of photobioreactors and the number of photobioreactorsused in conjunction with one another can vary greatly. Given theenormous scale required to manufacture industrially significant volumesof biofuels using photoautotrophic organisms, the dimensions of thephotobioreactors should be large. A large biofuel facility will cover athousand to many hundreds of thousands of acres. A preferred embodimentcomprises a width of about 2 to about 3 feet for the photobioreactor isthe average that would be optimal, but any size that incorporates theoverall design aspects of the photobioreactor technology can be used.The overall width and height of the photobioreactors can be determinedby evaluating a number of factors at the site of installation. Thedesired annual biofuels production volume, the desired cost and usefullife of the facility, the physical terrain of the facility site, thesolar exposure and the type of organisms that will be grown in thefacility are all considered when designing photobioreactors for aparticular location. A person skilled in the art of facility engineeringcan calculate the various cost/dimension relationship for thephotobioreactor tubes. The tube photobioreactors should be as long aspossible in order to minimize the number of connections in the system,which reduces the possibility of leaks that could lead to contamination.The tube photobioreactors can be constructed similar to the wayconventional pipelines which require a completely enclosed airtight orliquid tight environment are constructed. Recent advancements inplastics extrusion technologies allow extrusion of large-diameterplastic structures of over 6 feet in diameter. Extruded plastic elementscan make up the total structure of the photobioreactors. It is alsopossible to have an extruded lower part of the chamber made of rigidplastic and have the upper part of the chamber made of flexible plasticsheeting which is held in place by internal air pressure or supported byplastic or non-corrosive framing members at intervals. One can alsomanufacture the photobioreactor by molding, through any suitable moldingtechnique available including blow molding or rotational molding orextruding, in one or more parts and where necessary, then joining themolded units together.

Preferably, little or no external energy is needed to operate thephotobioreactors. In particular, it would be preferable if energy is notused to pump growth medium through the photobioreactors; however, insome instances it may be necessary to do so in order to provide churningand mixing to the growth medium, which ensures uniform exposure of cellsto solar radiation and promotes gas exchange between the growth mediumand the gas in the upper part of the chamber. Linear flow through solartubes can reduce the overall photosynthesis since cells near the middleor bottom of the tubes receive less light than those towards the top. Anembodiment hereof utilizes a flow of growth medium, preferably due togravity, and internal static mixers such as internal baffles to provideappropriate mixing. These flow baffles are static in nature, thusrequiring no additional energy input beyond the flow rate determined bythe pumping system and gravity. The careful balancing of mixing in thetubes with creating turbulent flow is important, since the cells aresensitive to sheer damage during heavy mixing. The design of thephotobioreactor in this embodiment mimics the mixing and churning in anatural stream lined with rocks. The turbulent flow in thephotobioreactor not only provides the necessary mixing for maximalphotosynthesis in high density cultures, but also serves to removedissolved oxygen and enhances absorption of CO₂ from the air/CO₂ mixturein the controlled air head above the culture. This also helps tostabilize the culture pH and increases the resident time of the biomassin the photobioreactor for photosynthesis.

The photobioreactor can also be fitted with dams of varying heights thatcan be used to control the flow of the culture in areas where there is agreater natural slope. This permits steeper surfaces to be used forproduction facilities and lower overall installation costs. In a furtherembodiment, the dams can provide the amount of churning needed forcultures such as Synechococcus or Synechocystis which grow in highdensities and need turbulence.

The dimensions of the photobioreactor provided herein can be varied topermit the mass culture of various genera of photoautotrophic organisms.In addition, the photobioreactor can accommodate different levels ofgrowth medium to meet the needs of various organisms. Preferably, thegrowth medium has a depth between about 6 inches and 12 inches.

A further embodiment of this method of ethanol collection and removal isto pass the gas phase or a portion of the gas phase through a cold trap,which results in the removal of essentially all of the ethanol from thegas phase. This portion of the gas phase can then be vented to the airto allow for removal of excess oxygen, or the oxygen can be collectedand used. Heat exchangers and the like can be used to reduce energycosts associated with cooling.

Although the present invention may be adapted to a vast number ofoverall physical configurations substantially similar to those shown inFIGS. 1-7, the concepts for growing genetically modifiedphotoautotrophic organisms in a closed system on a massive scale for theproduction of biofuels remain the same. The present invention overcomesthe well documented problems with open pond production systems, biomassgrowth and accumulation, as well as enables the cost effectiveconstruction of large capacity closed production systems. It will beunderstood by an ordinary person skilled in the art that the inventionwill not be limited to the specific designs and structures illustratedin the drawings.

FIG. 1 shows one photobioreactor of the present invention. Thephotobioreactor comprises an upper part of the chamber 1 and lower partof the chamber 2. Upper chamber 1 is at least partially translucent,preferably transparent. A temperature maintenance compartment 3 islocated below the lower part of the chamber to cool or heat the growthmedium 8. The photobioreactor may optionally have an interlocking tab 6to secure the photobioreactor to an adjoining photobioreactor, or asecuring tab 7 to fix the photobioreactor to a supporting surface. Oneor more supply inlets 5 add water, nutrients, carbon dioxide or anyother necessary inputs to the system. Ethanol, or other biofuel,released by genetically enhanced organisms in the growth medium 8evaporates into the headspace 9 in the upper part of the chamber 1 andcondenses on the inner surface 10 of the upper part of the chamber 1.

The photobioreactor depicted in FIG. 1 is in the shape of a rectangulartube; however other shapes and configurations are possible. FIG. 2 showsa photobioreactor in the shape of a hexagonal dome. The supply inlets 5can be positioned near the bottom of the lower part or any other part ofthe chamber 2. Additionally, the photobioreactor may contain a draintube 14, a mixing device 15 (in this embodiment an arm that is rotatedaround the photobioreactor) and a service tube 17 at the top of theupper part of the chamber 1 to provide access to the photobioreactor.The photobioreactor may also have condensate collection tube 16connected to the collection trough 4 inside the upper part of thechamber 1, and gas outlets 13. The gas outlets 13 are used to remove gasfrom the upper part of the chamber 1 to a gas collection and separationapparatus (not shown) and can serve as an oxygen exhaust valve to ventexcess oxygen out of the photobioreactor. FIG. 3 shows photobioreactorsof the present invention also having different designs.

FIGS. 4A-4B show a photobioreactor similar to that depicted in FIG. 1additionally containing multiple upper coolant compartments 18. Theupper coolant compartments 18 in FIG. 4A extend across the upper part ofthe chamber 1 and separate the upper part of the chamber 1 into multipleportions. In addition to promoting condensation of the biofuel bycooling the gas 9 in the upper part of the chamber 1, the upper coolantcompartments 18 provide increased inner surfaces 10 for condensation tooccur. The upper coolant compartment 18 can also be positioned along thesurface of the upper part of the chamber 1 as depicted in FIG. 4B.

FIG. 5 illustrates additional shapes and designs similar to thephotobioreactor of FIG. 1. The tube shape designs include circular tubesand other polygonal shapes. FIGS. 6 and 7 illustrate additionaldifferent designs of photobioreactor tubes suitable with the presentinvention.

In use, multiple photobioreactors can be placed next to one another FIG.8 and FIG. 9 to maximize production of ethanol of another biofuel. FIG.8 shows a three-dimensional end view of two photobioreactors of thepresent invention placed adjacent to one another. This configurationprovides easy access to the supply inlets 5. The condensed ethanol isremoved from the photobioreactor through condensate collection draintubes 16, which are connected to the collection trough 4 (not shown).Alternatively, a photobioreactor system is designed as severalphotobioreactors fused together as shown in FIG. 9.

Both ends of a tube shaped photobioreactor may have inlet and outlettubes, however, in some embodiments only one end may need to have inletsand outlets. FIG. 10 shows a rear view, side view, and front view of anend plug 20 that provides an air and watertight termination at the endsof photobioreactor. End plug 20 fits snugly into the ends, oralternately over the ends of the photobioreactor and provide connectionsto the supply inlets 5 and 19, drain tubes 14 and 16 and gas outlets 13.Also, caps can be installed over all inlets 5, 19 and outlets 13, 14 and16 to seal off any fittings. End plugs 12 can be permanently sealed tothe ends of photobioreactor or they can be held into place with variousfastening systems and sealed. By not permanently sealing the end plug12, access to the photobioreactor can be gained for repairs or cleaning.The connections can be of any size or shape that allow for the efficientexchange of air and inputs into the photobioreactor and can be straightpipe or threaded connections. The end plug 20 can be made as one singlepiece by molding or other acceptable means. End plugs 20 can beinstalled to a photobioreactor as shown in FIG. 11.

FIGS. 12 and 13 depict the front, rear and three-dimensional views ofthe photobioreactor flow connector 21. This flow connector 21 is used toconnect to the ends of two separate photobioreactors to continue theflow in the joined tubes in the opposite direction. Flow connector 21has openings on the top and on the back for access to thephotobioreactor for cleaning and repair. Flow connector 21 is connectedto the photobioreactors by inserting into the open ends of each andpermanently sealing the photobioreactors with appropriate adhesives. Thefinal design of flow connector 21 will be such that it will be able tojoin matching separate photobioreactors regardless of their finaldesign.

FIG. 14 depicts coupling unit 22 which is used to join a photobioreactorto another photobioreactor. FIG. 14 cross-section view shows the designof the coupling as well as how the coupling is attached to thephotobioreactor. The couplings should be permanently attached to twoseparate photobioreactors using adhesives. The installed coupling unit22 allows the connection of two short photobioreactor tubes into onelonger tube. FIG. 15 shows a three-dimensional view of an installedphotobioreactor coupling unit 22. The final design of coupling unit 28will be such that it will be able to join matching separatephotobioreactors regardless of their final design.

FIGS. 16-19 show various non-comprehensive configurations of the flowdams 24 and 28 and baffles 23, 25, 26, 27, 29, 30, 31, 32 and 33 pressedand/or molded into, or attached with adhesives to the inside bottom ofthe photobioreactor. FIGS. 16-19 do not embody all of the potentialvariations embodied in the invention.

FIG. 20 shows a three-dimensional view of various photobioreactorfittings. Various fittings 34, 35, 36 and 37 can be fixed tophotobioreactor by cutting the appropriate size hole in the body of thebioreactor and attaching the fittings using adhesives, solvents orsilicones. The fittings 34, 35, 36 and 37 can be used for venting ofexcess oxygen, addition or removal of various gases, nutrients, growthmedium, water and buffers as well as for various probes andenvironmental monitoring devices. Each fitting can contain a valve 37,either electronic or manual, to control when the fitting is open orclosed.

In situations where the growth medium is pumped or otherwise caused toflow through the photobioreactor, the present invention provides severalfeatures which provide an optimal environment for organism growth. Inparticular, mixing or churning ensures that the cells in the culture aremore evenly exposed to sunlight, releases excess dissolved oxygen fromthe growth medium, and improves absorption of carbon dioxide. Thephotobioreactor shown in FIG. 21A is placed on a slope and utilizesgravity to transport the growth medium 8 through the lower part of thechamber 2. Alternatively, the growth medium 8 is pumped through thelower part of the chamber 2, as shown in FIG. 21B. As shown in FIGS. 21Aand 21 B, the bottom surface of the lower part of the chamber 2 containsvarious integrated flow dams 24 and 28 and baffles 23, 25, 26, 27, 29,30, 31 and 32 to provide significant churning in the growth medium 8.The flow dams 24 and 28 and baffles 23, 25, 26, 27, 29, 30, 31, 32 and33 may also have the configurations depicted in FIGS. 16-19. These flowdams 24 and 28 and baffles 23, 25, 26, 27, 29, 30, 31 and 32 can beintegrated into or attached to the bottom surface of the lower part ofthe chamber 2. A person skilled in the art of fluid dynamics can readilydesign the optimal configuration of the flow dams 24 and 28 and baffles23, 25, 26, 27, 29, 30, 31 and 32 based on the variables of the plantlocation and needs of the organisms.

As illustrated in FIG. 22, sunlight enters through the top of the upperpart of the chamber 1. Algae in the growth medium 8 uses the sunlight toproduce and release ethanol into the growth medium where it isevaporated 9 into the upper part of the chamber 1. The gas 9 in theupper part of the chamber will include oxygen, evaporated water andethanol. Ethanol will condense on the inner surface 10 of the upper partof the chamber 1, and the condensate 11 will collect into the condensatecollection trough 4. As depicted in FIG. 22, carbon dioxide is providedto the growth medium 8 as a gas present in the lower part of the chamber1. Thus, the growth medium 8 absorbs carbon dioxide and releases oxygeninto the upper part of the chamber 1. Various inlets 5 and 19 supply thenecessary inputs into the system and outlets remove 13, 14 and 16 removegases, liquid and biomass from the system.

The biological processes in the present invention do not produce ethanolin the presence of water and other materials or other biofuel product.Rather, the product must be purified to separate it from water, andpossibly other cellular products and contaminants. Therefore anadditional aspect of the production of ethanol with a photobioreactor isthe purification of ethanol or other biofuels. The biofuel product canbe purified from the gas phase of the photobioreactor as well as fromthe condensates. The elimination of oxygen from a biomass producingphotobioreactor is generally accomplished by venting a gas phaseenriched in oxygen to the atmosphere. While this same method can be usedwith the present invention, the oxygen stream will contain the biofuelproduct and may substantially reduce the productivity of thephotobioreactor unless measures are taken to prevent excessive productloss. Therefore another aspect of the present invention is the controlof biofuel loss, particularly ethanol loss, in the oxygen exhauststream.

The condensate can be removed as a liquid and purified by any knownmeans, such as distillation, pervaporation, dephlegmation, etc., and maybe further dehydrated by any known means such as zeolite dehydration ormembrane dehydration.

Integration of Oxygen Exhaust and Ethanol Harvest Processes

The removal of a substantial amount of ethanol in the gas phase isunavoidable due to the requirements for oxygen removal. At a minimum, itis necessary to avoid losing too much ethanol as a consequence of oxygenrelease. Ethanol retention during oxygen release is a separationprocess, and it may be that the release of oxygen and the separation ofethanol from the vapor stream can be combined into a single operationthat is more cost effective than independent operations would be. Forexample, the gas phase from the photobioreactor could be pumped past anoxygen permeable membrane at an elevated pressure and allowed toequilibrate with air through the membrane. The partial pressure ofoxygen in the output gas from such a system would be over 21 kPa by anamount dependent on how closely equilibrium is approached. The remainderof the vapor phase would be ethanol and water. The vapor phase couldfurther be passed through a dephlegmator to remove most of the water,and through a dehydration system to remove most of the remaining water.

Exclusion of Heterotrophs

Because ethanol is not always stable to biological degradation in thepresence of oxygen, it is important to exclude, inhibit or killheterotrophic organisms so that degradation of ethanol by heterotrophsis minimal. Methods for sterilization of equipment, solutions and gassesare well known to those skilled in the art, as is the preparation andmaintenance of cyanobacterial or algal cultures which are axenic, thatis, that contain no organisms of any kind other than the cyanobacterialor algal strain being cultured. Chemical treatments such as antibioticsare also well known, as are techniques for making cell lines ofcyanobacteria or algae that are resistant to particular antibiotics.Antibiotics may be added to the cultures in the photobioreactors toeliminate any heterotrophic organisms present in the culture or as aprophylactic measure against potential infection. Additional methods forthe control of heterotrophs as known in the art may also proveeffective. The preferred embodiment of the invention is that thecyanobacteria or algae organisms themselves produce ethanol in thephotobioreactor and when this level is near or above 5% in the culture,the ethanol itself will kill heterotrophs and assist in keeping theculture stable and near sterile.

Having now fully described the photobioreactors and methods providedherein in some detail by way of illustration and examples for purposesof clarity of understanding, it will be obvious to one of ordinary skillin the art that the modifying or changing the photobioreactorconfigurations and methods to utilize equivalent conditions, elements,steps, and other parameters is within the scope of the appended claims.

One of ordinary skill in the art will appreciate that startingmaterials, substrates, device elements, analytical methods, mixtures andcombinations of components other than those specifically exemplified canbe employed in the practice of the invention without resort to undueexperimentation. All art-known functional equivalents, of any suchmaterials and methods are included in the devices and methods disclosedherein. The terms and expressions which have been employed are used asterms of description and not of limitation, and there is no intention inthe use of such terms and expressions of excluding any equivalents ofthe features shown and described or portions thereof, but it isrecognized that various modifications are possible within the scope ofthe claims. Thus, it should be understood that although thephotobioreactors and methods provided herein have been specificallydisclosed in terms of embodiments, optional features, modifications andvariation of the concepts herein disclosed can be utilized by thoseskilled in the art, and such modifications and variations are consideredto be within the scope of the appended claims.

As used herein, “comprising” is synonymous with “including,” or“containing,” and is inclusive or open-ended and does not excludeadditional, unrecited elements or method steps.

When a group of materials, compositions, components or compounds isdisclosed herein, it is understood that all individual members of thosegroups and all subgroups thereof are disclosed separately. When aMarkush group or other grouping is used herein, all individual membersof the group and all combinations and subcombinations possible of thegroup are intended to be individually included in the disclosure. Everyformulation or combination of components described or exemplified hereincan be used to practice the invention unless otherwise stated. Whenevera range is given in the specification, for example, a temperature range,a time range, or a composition range, all intermediate ranges andsubranges, as well as all individual values included in the ranges givenare intended to be included in the disclosure. In the disclosure and theclaims, “and/or” means additionally or alternatively. Moreover, any useof a term in the singular also encompasses plural forms.

All references cited herein are hereby incorporated by reference intheir entirety to the extent that there is no inconsistency with thedisclosure of this specification. Some references provided herein areincorporated by reference to provide details concerning sources ofstarting materials, additional starting materials, additional reagents,additional methods of analysis, additional biological materials,additional cells, and additional uses of the invention. All headingsused herein are for convenience only. All patents and publicationsmentioned in the specification are indicative of the levels of skill ofthose skilled in the art to which the invention pertains, and are hereinincorporated by reference to the same extent as if each individualpublication, patent or patent application were specifically andindividually indicated to be incorporated by reference. References citedherein are incorporated by reference herein in their entirety toindicate the state of the art as of their publication or filing date andit is intended that this information can be employed herein, if needed,to exclude specific embodiments that are in the prior art.

The meaning of terms includes text as follows.

The term “closed from the outside environment” includes the meaning thatbacteria outside of the photobioreactor are restricted from enteringinto the photobioreactor and that ethanol formed within thephotobioreactor is restricted from leaving the photobioreactor otherthan in predesigned ways.

The term “chamber” includes the meaning of the body of thephotobioreactor which comprises an aqueous growth medium comprising aculture of genetically enhanced organisms disposed in the growth medium,wherein said organisms are selected from the group consisting of algaeand cyanobacteria and a means to furnish sunlight to the organisms.

There is an upper part of the chamber which comprises a translucent orclear region to allow in sunlight. There is a lower part of the chamberwhich comprises an aqueous growth medium. The chamber encloses agas-phase volume or head space. The chamber encloses a liquid-phasevolume which comprises the aqueous growth medium. There is an innersurface of the upper part of the chamber upon which gas-phase water andethanol can condense. The meaning of the term chamber may also beunderstood from the Figures.

The term “upper part of the chamber” or “upper chamber” comprises theregion of the chamber which is translucent or clear.

The term “lower part of the chamber” or “lower chamber” comprises theregion of the chamber comprising the aqueous growth medium. No physicalbarriers between the upper parts of the chamber and the lower parts ofthe chamber are required.

The term “continued daily basis” includes the meaning of the operationof the photobioreactor and refers to the continued production of ethanolfrom carbon dioxide by the organisms during the continued times forwhich sufficient light is available. The term “continued daily basis”further includes the meaning that ethanol can be formed day after dayprovided that there is sufficient light, carbon dioxide and nutrientspresent. The term “continued daily basis” further includes the meaningthat the production of ethanol will be influenced by the weather,including the presence or absence of sun or clouds, and by the need toinclude nutrients, promoters, and carbon dioxide in the chamber, and byroutine maintenance requirements, including the need to replace biomassand/or components of the photobioreactor.

The term “inner surface of the upper part of the chamber” includes theinner portion of the chamber upon which gas-phase ethanol and watercondense to the liquid phase.

The term “inner surface of the upper part of the chamber” includes theinner portion of the chamber upon which gas-phase ethanol and watercondense to the liquid phase.

The term “inlet and outlet tubes” refers to tubes connected to openingsin the chamber of the photobioreactor.

The term “coolant compartments” refers to compartments that holdmaterial that may be used to cool the chamber of the photobioreactor.

The term “mixing device” refers to any device that may be used to mix orstir the contents of the aqueous growth medium.

The term “measuring devices” refers to devices that may makemeasurements of parameters related to the photobioreactor and includesdevices to measure temperature, pressure, and the amount of carbondioxide, carbonate, salt, hydrogen ion, water, ethanol, oxygen andnutrient levels of the growth medium.

The term “treatment of the inside of the upper part of the chamber”includes any chemical or physical treatment of the surface of the insideof the upper part of the chamber of the photobioreactor which can affectthe condensation of gas phase ethanol.

The term “additive, coating and physical modifications” includes anychemical or physical modifications of the chamber of the photobioreactorthat can affect the photosynthesis reaction or the condensation ofethanol or water inside the chamber. Additives can include UV blockingagent, UV stabilizing agent, UV blocking coating, UV blocking film, anadditive for stabilizing the plastic material, and agents forstabilizing the plastic material.

The term “closed reaction volume” includes the meaning that the volumedefined by chamber of the photobioreactor is restricted to the entry ofentities detrimental to the photosynthetic reaction and restricted as tothe departure of ethanol from the reaction volume. The term “closedreaction volume” does not indicate that oxygen does not leave thereaction volume or that ethanol does not leave the reaction volume.

The term “liquid phase” includes both the liquid in the culture, and theliquid as a condensate, and includes both H2O and ethanol liquids.

The term “modular plastic extrusions, mounted on soil” includes themeaning of a plurality of photobioreactor units, comprising modularplastic, mounted on soil.

The term “means of temperature regulating the aqueous growth medium”refers to a means of temperature regulation which includes structureswhich regulate temperature through variation of light input or variationof temperature by input of heated or cooled fluids. The temperature ofthe aqueous growth medium can be regulated by electric heaters orthermal contact with heated or cooled fluids.

The means of introducing fluids and gases could include a nozzle, avalve, and venturi connected to a source of CO₂ which could include fluegas from a power plant of CO₂ from a cylinder

The means of introducing carbon dioxide can include introduction througha tube from a CO₂ cylinder or from combustion gases of a fossil fuelcombustion unit.

The means of converting the carbon dioxide into ethanol includephotosynthetic processes involving microorganisms selected from thegroup consisting of genetically enhanced algae and cyanobacteria.Genetically enhanced algae and cyanobacteria (as described for examplein U.S. Pat. No. 6,699,696) convert carbon dioxide into ethanol.

The means of separating ethanol from the liquid phase include astructure which allows for evaporation of ethanol from an aqueous growthmedium into a headspace and subsequent condensation onto the surface ofa chamber. The structure may include a semipermeable membrane.

An important discovery of this invention is that for photobioreactors ofthis invention is that solar evaporation allows for an enhancement inethanol concentration. For a given concentration X of ethanol in theliquid-phase including the microorganisms we have data suggesting thatthe condensate formed by solar evaporation of the same liquid-phase isgreater than X. This is based on data for the evaporation of ethanol andwater from a liquid of ethanol and water in a prototype photobioreactoras illustrated below, with the data denoted (“Photobioreactor”)corresponding to liquid in the lower part of the bioreactor chamber(“Pond”) and with the data denoted (“Condensate”) corresponding to the“evaporated then condensed” liquid. Photobioreactor Apparatus #5 andPhotobioreactor Apparatus #6 are two example photobioreactors whichembody the invention illustrate the enrichment or enhanced ethanolconcentration.

Data for Photobioreactor Apparatus #5

Ethanol Sample Ethanol Sample concentration volume in sample Day From(mM) (L) (mmol) 1 Pond 224.04 1 Condensate 279.23 1.48 411.87 2 Pond224.99 2 Condensate 275.83 2.00 551.67 3 Pond 222.37 3 Condensate 254.800.98 248.43 6 Pond 221.61 6 Condensate 256.30 4.10 1050.82 7 Pond 215.267 Condensate 273.83 0.90 246.45 8 Pond 212.93 8 Condensate 247.37 1.28315.39 9 Pond 216.86 9 Condensate 253.06 0.73 183.47 10 Pond 253.93 10Condensate 210.78 1.30 274.01 13 Condensate 169.57 3.80 644.37 14Condensate 158.60 1.50 237.90 15 Pond 132.35 15 Condensate 166.46 1.43237.21 Sum of condensate 19.48 4401.60The ethanol yield in the condensate is 4.4 moles=203 g. Over 15 days theyield is 13.5 g/day, which for the active surface area of 1.42 m2 is 9.5g/(d-m2).

The enrichment in ethanol, condensate/pond, for day 1 is 1.25.

Data for Photobioreactor Apparatus #6

1 Pond 230.84 1 Condensate 289.87 1.225 355.1 2 Pond 231.89 2 Condensate292.67 1.6 468.3 3 Pond 227.12 3 Condensate 267.81 0.675 180.8 6 Pond228.27 6 Condensate 270.83 3.725 695.6 7 Pond 225.92 7 Condensate 273.390.75 205.0 8 Pond 229.38 8 Condensate 273.83 1.1 301.2 9 Pond 232.04 9Condensate 280.41 0.575 161.2 10 Pond 233.75 10 Condensate 272.41 1.175320.1 13 Condensate 186.75 2.9 541.6 14 Condensate 197.80 1.225 242.3 15Pond 159.70 15 Condensate 192.10 1.2 230.5 Sum of condensate 16.153701.8

In a different reactor, an experiment was done with an ethanol solutionat an initial level of 0.5% (v/v). The water phase comprised salt.Specifically, seawater was added to the reactor to an original depth of˜15.24 cm (6 in). The liquid volume was about 50 liters. The initialsalinity of the water was 35 ppt. In this case, the ratio of ethanolconcentration for condensate/pond was of the order of 3, higher than the1.25 mentioned above. Salt is believed beneficial to enhanced ratios ofethanol/water in the condensate. In a working photobioreactor, the levelof salt must be compatible with the functioning of the ethanol-producingcyanobacteria. In a working photobioreactor, the temperature must becompatible with the functioning of the ethanol-producing cyanobacteria.A preferred temperature range is 20 degree Celsius to 60 degree Celsius.

Example of a Flexible Tube Bioreactor

We constructed a 4.45′×50′ tube photobioreactor with condensatecollection troughs.

We special ordered (AT Films, Inc., Edmonton, Alberta) 10,000 pounds ofhigh-density polyethylene (HDPE) 6 mil tubular film 43.7 inch diameterthat was UV stabilized to last 4 years and included an anti-fogadditive. The anti-fog additive reduces the light blocking of condensatecollected on the surface of the photobioreactor from ˜20% to ˜4%. Thetubular film was delivered on 500′ rolls, and the film has a lay flatwidth of 5.708′. Starting with 50 feet of 6 mil tubular film laid flaton a long table we marked two lines the length of the tube 2.75″ in fromeach edge.

This is our proposed water line when the photobioreactor is built. Thefinal shape of the tube photobioreactor can be calculated by a personskilled in the art of mechanical engineering. We wanted our finalphotobioreactor tube width to be ˜53″ with 8″ of water. Our mechanicalengineer provided the shape of the photobioreactor tube (see figure) andthe location of the waterline.

We start on one side of the tube and pushed in a fold “finger” to theinside of the tube (see figure). We adjusted the length of the finger tothe final desired width of the collection trough 3″. We folded thisfinger to the inside of the tube the whole length of the tube. We madesure the fold is even with the water line mark on the bag. We used threelong pieces (aluminum—20′×1.74″×1.75″) of angle iron to clamp the tubeto the edge of the table, so 0.5″ of the finger fold is hanging over theedge of the table and is held in place with the angle irons which areclamped on each end.

This is to hold the fold and the tube firm so the heat sealer can be rundown the length of the tube without the tube moving. This “tongue” is 4layers of the 6 mil tube with the water line mark about 0.5″ from theedge of the table. Using a Bosch HS-CII portable/tabletop compactcontinuous band sealer (BaggingGuys.com) with the heat setting on 3.5(˜400 F), we sealed the finger the length of the tube.

We repeated this on the other side of the tube. The circumference of thefinished tube was 124″. We special ordered two fiberglass (RileyComposites, Inc., West Palm Beach, Fla.) end caps which will fit insidethe tube to provide a means to seal the end and support the tubingfittings for water and air inlets/outlets (whatever size is desired, weused 0.5″ BKF124, Aquatic Ecosystems, Inc., Apopka, Fla.).

The end caps have a shape like the tube reactor will have when it hasthe desired water level (8″) and air pressure (0.6 psi). We marked thedepth of the end cap from each end of the tube. We marked a point 4″from the inside of the end cap above the condensate collection trough.With a 0.5″ hole saw, we cut a 0.5″ round hole through the plastic andinstalled a 0.5″ bulkhead fitting (0.5″ BKF124, Aquatic Ecosystems,Inc., Apopka, Fla.) with gasket. We did this on one end of the tube (theexpected downslope side, if the final installation has a slope. If noslope is expected, then either or both ends can have the outlets).

We then installed 0.5″ FIPT/⅜″ barbed nylon elbows (The Home Depot) inthe outside of the bulkhead fittings. With a 0.5″ hole saw, we cut two0.5″ round holes through the end cap, one above the water line and onebelow the waterline and installed 0.5″ bulkhead fitting (0.5″ BKF124,Aquatic Ecosystems, Inc., Apopka, Fla.) with gasket. We then installed0.5″ FIPT/FIPT nipples (The Home Depot) in the outside of the bulkheadfittings and attached plastic 0.5″ FIPT/FIPT ball valves to the nipples.We then installed a ½″× 3/16″ butyl tape gasket (Glue Products, Inc.,West Palm Beach, Fla.) around the end cap and pulled the tube over thegasket.

We pressed the tube to the gasket. We used a ½″ Manual CombinationStrapping Tool and ½″ Machine Grade Polypropylene Strapping(GlobalIndustrial.com) to apply a strap around the outside of the endcap over the tube and butyl tape and applied sufficient torque to squishthe butyl tape to create a seal between the end cap and the tube.

At the opposite end of the bioreactor, we stuck a compressed air line inthe open end of the fingers, and pumped compressed air into the insideof the finger for floatation 100. We completed the same sealing processon this end. The completed tube was filled with water to the bottom ofthe condensate collection troughs and air was pumped into the tube untilit was taught. Inside the completed tube the water evaporates andcondenses on the upper inner surface of the photobioreactor, flowing tothe condensate collection trough.

Once in the condensate collection trough, the water flows down thetrough via gravity and out the bulkhead fitting for collection.

The photobioreactor formed in the above-example can be implemented inthe following way.

A photobioreactor system formed by joining an approximately cylindricalannulus of flexible, visible-light transmitting, plastic sheet withrigid endcaps at each end of the annulus to form a closed chamber,further comprising

a. a plurality of openings within one or more of the endcaps for inletand outlet tubes;

b. a lower part of the chamber which comprises an aqueous growth mediumcomprising a culture of genetically enhanced organisms disposed in thegrowth medium, wherein said organisms are selected from the groupconsisting of algae and cyanobacteria, and wherein said organismsproduce ethanol on a continued daily basis which enters the growthmedium;

c. an upper part of the chamber, comprising a headspace above theaqueous growth medium, wherein the headspace comprises carbon dioxide;

d. an inner surface on the upper part of the chamber upon which theethanol formed by said organisms condenses; and

e. a plurality of troughs for the collection of the condensed ethanol

In other embodiments, the closing of the ends of the cylindrical annuluscan be accomplished without the use of rigid endcaps.

Photobioreactor Apparatus # 5 Photobioreactor Apparatus # 6 Sample DateSample Time Sample Type Ethanol (mM) Sample Date Sample Time Sample TypeEthanol (mM) Day 1 9:00 AM Photobioreactor 222.04 Day 1 9:00 AMPhotobioreactor 23O.84 Day 1 9:00 AM Condensate 279.23 Day 1 9:00 AMCondensate 289.87 Day 2 9:30 AM Photobioreactor 224.99 Day 2 9:30 AMPhotobioreactor 231.89 Day 2 9:30 AM Condensate 275.83 Day 2 9:30 AMCondensate 292.67 Day 3 9:00 AM Photobioreactor 222.37 Day 3 9:00 AMPhotobioreactor 227.12 Day 3 9:00 AM Condensate 254.80 Day 3 9:00 AMCondensate 267.81 Day 6 10:00 AM  Photobioreactor 221.61 Day 6 10:00 AM Photobioreactor 228.27 Day 6 10:00 AM  Condensate 256.30 Day 6 10:00 AM Condensate 270.83 Day 7 9:00 AM Photobioreactor 215.26 Day 7 9:00 AMPhotobioreactor 225.92 Day 7 9:00 AM Condensate 273.83 Day 7 9:00 AMCondensate 273.39 Day 8 9:00 AM Photobioreactor 212.93 Day 8 9:00 AMPhotobioreactor 229.38 Day 8 9:00 AM Condensate 247.37 Day 8 9:00 AMCondensate 273.83 Day 9 9:00 AM Photobioreactor 216.86 Day 9 9:00 AMPhotobioreactor 232.04 Day 9 9:00 AM Condensate 253.06 Day 9 9:00 AMCondensate 280.41 Day 10 9:00 AM Photobioreactor 210.78 Day 10 9:00 AMPhotobioreactor 233.75 Day 10 9:00 AM Condensate 253.93 Day 10 9:00 AMCondensate 272.41

1. A photobioreactor system formed by joining an approximatelycylindrical annulus of flexible, visible-light transmitting, plasticsheet to form a closed chamber, further comprising a. a plurality ofopenings formed in the chamber for inlet and outlet tubes; b. a lowerpart of the chamber which comprises an aqueous growth medium comprisinga culture of genetically enhanced organisms disposed in the growthmedium, wherein said organisms are selected from the group consisting ofalgae and cyanobacteria, and wherein said organisms produce ethanol on acontinued daily basis which enters the growth medium; c. an upper partof the chamber, comprising a headspace above the aqueous growth medium,wherein the headspace comprises carbon dioxide; d. an inner surface onthe upper part of the chamber upon which the ethanol formed by saidorganisms condenses; and e. a plurality of troughs for the collection ofthe condensed ethanol, wherein the troughs are supported by a flotationdevice in contact with the aqueous growth medium.
 2. The photobioreactorsystem of claim 1, wherein the troughs are formed from the flexiblelight transmitting plastic sheet.
 3. The photobioreactor system of claim1, wherein the troughs are formed from the flexible light transmittingplastic sheet, wherein said troughs are supported by air added to thevoid formed in the construction of each trough to serve as the flotationdevice in contact with the aqueous growth medium.